ДИФФЕРЕНЦИРОВАНИЕ ПЛЮРИПОТЕНТНЫХ СТВОЛОВЫХ КЛЕТОК

... 1. Способ повышения выхода клеток, экспрессирующих маркеры, характерные для линии сформированной эндодермы, путем дифференцирования плюрипотентных стволовых клеток в клетки, экспрессирующие маркеры, характерные для линии сформированной эндодермы, где способ включает обработку полипотентных стволовых клеток средой, в которой отсутствует активин А и содержится GDF-8 и анилин-пиридинотриазин, где анилин-пиридинотриазин представляет собой 14-проп-2-ен-1-ил-3,5,7,14,17,23,27-гептаазатетрацикло[19.3.1.1~2,6~.1~8,12~]гептакоза-1(25),2(27),3,5,8(26),9,11,21,23-нонаен-16-он, в течение периода времени, достаточного для того, чтобы плюрипотентные стволовые клетки дифференцировались в клетки, экспрессирующие маркеры, характерные для линии сформированной эндодермы.2. Способ по п. 1, в котором способ повышает процент клеток, экспрессирующих CXR4.3. Способ дифференцировки плюрипотентных стволовых клеток в клетки, экспрессирующие маркеры, характерные для линии сформированной эндодермы, включающий обработку ...

BIOLOGISCHE KÜNSTLICHE LEBER

CELL AND VIRUS CULTURE SYSTEMS

... 1436323 Preparing vaccines WELLCOME FOUNDATION Ltd 25 April 1973 [26 April 1972] 19387/72 Heading A5B [Also in Division C6] A vaccine may be prepared from a virus, e.g. foot and mouth disease virus, which has been cultured within a porous carrier which consists of pairticulate material which retains the virus whilst allowing liquid media, e.g. nutrient, to pass through. The inactivated virus is formulated into a vaccine in known manner.

Development of spinal cord on a microfludic chip

Novel Microcarrier beads

A microcarrier bead made from apatite characterised by one or more of the following: (a) micro-metre sized; (b) regular porous structure; (c) rough surface; (d) substantially spherical; (e) osteo-conductivity; (f) chemical similarity to the mineral phase of bone; and (g) high thermal stability. Preferably the apatite is selected from: hydroxyapatite, silicon-substituted apatite, silver-substituted apatite, magnesium-substituted apatite and a stoichiometric apatite which is a synthetic apatite with a Ca/P atomic ratio that approaches 1.67. Preferably the apatite is phase-pure and the beads are between 100-800µm diameter. A method for making microcarrier beads is outlined comprising (a) mixing apatite and alginate in a solution and allowing them to disperse to form a suspension; (b) extruding the suspension dropwise through a droplet device; (c) exposing the extruded droplets to a calcium chloride solution and dispersing the same; (e) hardening the beads in a solution of alcohol; (f) drying ...

IN-VITRO PRODUCTION OF TRANSPLANTIERBAREM CARTILAGE FABRIC

PROCEDURE FOR THE PRODUCTION OF VIRUS MATERIAL

COKULTUR LTE (LYMPHOID TISSUE EQUIVALENT) FOR ARTIFIZIELLES IMMUNE SYSTEM (AIS)

PROCEDURE FOR THE PRODUCTION OF FSME VIRUS ANTIGEN

REKOMBINANTER ZELLKLON MIT ERHÖHTER STABILITÄT IN SERUM- UND PROTEINFREIEM MEDIUM UND VERFAHREN ZUR GEWINNUNG DES STABILEN ZELLKLONS

IN-VITRO PRODUCTION OF TRANSPLANTIERBAREM CARTILAGE FABRIC

BIOTHERAPEUTIC MICRO SPHERES COATS WITH CELLS

Development of spinal cord on a microfluidic chip

The invention relates to culturing brain endothelial cells, and optionally astrocytes and neurons in a fluidic device under conditions whereby the cells mimic the structure and function of the blood brain barrier. Culture of such cells in a microfluidic device, whether alone or in combination with other cells, drives maturation and/or differentiation further than existing systems.

Methods for cell expansion and uses of cells and conditioned media produced thereby for therapy

A method of cell expansion is provided. The method comprising culturing adherent cells from placenta or adipose tissue under three-dimensional culturing conditions, which support cell expansion.

Automatable artificial immune system (AIS)

Three-dimensional peripheral lymphoid organ cell cultures

Scalable process for cultivating undifferentiated stem cells in suspension

Tissue compositions using cultured fibroblasts and keratinocytes and methods of use thereof

In vitro production of transplantable cartilage tissue

CELL THERAPY FORMULATION METHOD AND COMPOSITION

Ex-vivo prepared T-cells are harvested from cell culture conditions and formulated in medium suitable for infusion. The formulation is made by labeling the cells with one or more agents which have reactivity for T-cell surface moieties capable of delivery activation signals upon cross-linking and mixing the labeled cells with biodegradable nanospheres or microspheres coated with a material capable of cross-linking the agents attached to the T-cell surface moieties. Alternatively, the formulation may be made by mixing a population of T-cells with biodegradable nanospheres or microspheres coated with a first material and one or more second materials. The first material binds the second material and the second material has reactivity for surface moieties on the T-cells and the interaction of the second materials with the T- cells causes the activation of the T-cells. In either method, the mixture of T- cells and biodegradable spheres are suspended in a medium suitable for infusion, and the ...

MICROCARRIER CELL CULTURE REACTOR SYSTEM

Anchorage-dependent cells are grown in agitated microcarrier suspension in which the cells and microcarriers are aggregated by periodically providing a temporary residence of said microcarriers and cells outside the main cell culture reactor agitation zone and in a separate compartment wherein they are subjected to a gentle tumbling action within a confined space having substantially the same environmental conditions as in the main cell culture reactor.

PROCESSES FOR GENERATING ENGINEERED CELLS AND COMPOSITIONS THEREOF

The present disclosure provides processes for genetically engineering T cells, such as primary CD4+ T cells and/or CD8+ T cells, for use in cell therapy that does not involve expanding the cells. In particular aspects, the provided processes successfully generate compositions of engineered T cells, such as containing populations of engineered T cells, that express a chimeric antigen receptor (CAR) within a shortened amount of time as compared to alternative engineering processes, such as processes that involve expanding the cells. In certain aspects, the provided processes successfully generate a composition of engineered T cells suitable for use in cell therapy within 4 days from when the process to stimulate or activate the cells is initiated. In some aspects, the resulting engineered cell compositions are composed of cell population that are less differentiated, less exhausted, and more potent than engineered T cell compositions generated by other means, such as by processes that involve ...

NEUROMUSCULAR JUNCTION

The invention relates to culturing motor neuron cells together with skeletal muscle cells in a fluidic device under conditions whereby the interaction of these cells mimic the structure and function of the neuromuscular junction (NMJ) providing a NMJ-on-chip. Good viability, formation of myo-fibers and function of skeletal muscle cells on fluidic chips allow for measurements of muscle cell contractions. Embodiments of motor neurons co-cultures with contractile myo-fibers are contemplated for use with modeling diseases affecting NMJ's, e.g. Amyotrophic lateral sclerosis (ALS).

IN VITRO EXPANSION OF POSTPARTUM-DERIVED CELLS USING MICROCARRIERS

Compositions and methods for the growth and expansion of mammalian cells in culture are provided. In particular, methods for the growth and expansion of postpartum-derived cells in vitro are provided using surfaces such as microcarrier beads.

AUTOMATABLE ARTIFICIAL IMMUNE SYSTEM (AIS)

SCALABLE PROCESS FOR CULTIVATING UNDIFFERENTIATED STEM CELLS IN SUSPENSION

... ²²²The invention relates to a process for cultivating undifferentiated stem cells ²in suspension and in particular to a method for cultivating stem cells on ²microcarriers in vessels. The method enables stem cells to be cultivated in a ²scalable process for the first time, without losing their pluripotentiality. ²Different types of vessels are suitable for cultivation, such as e.g. spinners ²and bioreactors. The option of precisely setting the cultivation conditions in ²said vessels aids the process of obtaining a pluripotentiality of the stem ²cells. The inventive method also permits large yields of stem cells of an ²unvarying quality to be produced.² ...

TEMPERATURE-RESPONSIVE MICROCARRIER

The invention relates to compositions and methods useful for cell culture in which cell adherence and release of cultured cells can be performed in a temperature-responsive manner.

SUSPENSION AND CLUSTERING OF HUMAN PLURIPOTENT STEM CELLS FOR DIFFERENTIATION INTO PANCREATIC ENDOCRINE CELLS

The present invention provides methods of preparing aggregated pluripotent stem cell clusters for differentiation. Specifically, the invention discloses methods of differentiating pluripotent cells into beta cell, cardiac cell and neuronal cell lineages using suspension clustering. The methods involve preparing the aggregated cell clusters followed by differentiation of these clusters.

CULTURE METHOD AND CELL CLUSTER

The present invention is a culture method for culturing a population consisting of two or more cells including stem-cell-derived cells and mesenchymal cells in an indentation (10). Stem-cell-derived cells are cells obtained by differentiating stem cells in vitro. Such cells are one or more types of cell selected from the group consisting of endodermal cells, ectodermal cells, and mesodermal cells. The population is cultured in an indentation (10) together with vascular cells or secretory components. The indentation (10) has a space in which the cells can move. The volume of the space is taken to be V mm3. The number of mesenchymal cells seeded in the space is taken to be N. At this time, V is 400 or less. Furthermore, N/V is from 35 to 3000.

A METHOD FOR TRANSPLANTING CELLS INTO THE BRAIN AND THERAPEUTIC USES THEREFOR

... 2128630 9314790 PCTABS00024 A method for grafting a cell in the brain of a mammalian subject is accomplished by attaching the cell to a support matrix so that the cell attaches to the matrix surface, and implanting the support matrix with the attached cell into the brain. Preferred support matrices are glass or plastic microbeads, either solid or porous, having a diameter from about 90 to about 125 .mu.m. The method employs cells of different types, preferably cells of neural or paraneural origin, such as adrenal chromaffin cells. Also useful are cell lines grown in vitro. Cells not of neural or paraneural origin, such as fibroblasts, may also be used following genetic alteration to express a desired neural product such as a neurotransmitter or a neuronal growth factor. The method is used to treat neurological diseases such as Parkinson's disease, Alzheimer's disease, Huntington's disease, epilepsy, and traumatic brain injury.

BIOLOGICAL ARTIFICIAL LIVER

... 2107582 9316171 PCTABS00025 Disclosed is a hepatocyte bioreactor or bio-artifical liver which comprises a containment vessel having a perfusion inlet means and a perfusion outlet means; a matrix within said containment vessel so as to entrap hepatocyte aggregates within said containment vessel while allowing perfusion of said matrix. The invention is useful as an artificial liver.

BIOTHERAPEUTIC CELL-COATED MICROSPHERES

A living skin replacement for the treatment of partial-and full-thickness skin i njuries, such as burns and other wounds, consists of a slurry of cell-coated microspheres which can be applied to the ski n injury in much the same manner as a paste or salve. The skin implant can accommodate contour variations across the often exte nsive area of a skin injury and does not require the use of stapling, suturing, or other attachment methods. The microspheres can be formed of a variety of materials that are bio-compatible and resorbable in vivo.

MATRIX WITH ADHERENTLY BOUND CELLS AND PROCESS FOR PRODUCING VIRUSES/VIRUS ANTIGENS

Procédé de culture de cellules

Process for the cultivation of anchorage-dependent cells in a microcarrier culture, microcarriers for carrying it out and use of the process.

IMPROVED PREPARATIONS CELLS - PRECURSORS OF MATURE LIVER

A METHOD FOR PRODUCING A HETEROLOGOUS SECRETED PROTEIN FROM CHINESE HAMSTER OVARIES CELLS GROWN ON MICROCARRIERS

Neural networks formed from cells derived from pluripotent stem cells

POROUS ORGANIC/INORGANIC COMPOSITE AND MANUFACTURING METHOD THEREOF

The present invention relates to a porous organic/inorganic composite and a manufacturing method thereof and, more specifically, to a porous organic/inorganic composite in which a plurality of macropores is three-dimensionally connected to each other to form a plurality of paths and a manufacturing method thereof. The porous organic/inorganic composite according to the present invention has a structure in which a plurality of macropores is connected to each other, thereby being usefully used as a cell culture support and applied in various fields such as a catalyst, an absorbent, a sensor, a drug carrier, etc. COPYRIGHT KIPO 2017 (AA) Pores (BB) Skeleton (CC) Path ...

Culture of Pluripotent and Multipotent Cells on Microcarriers

Novel Microcarrier Beads

Novel Microcarrier 5 The invention relates to a novel microcarrier bead; a method for producing same; a therapeutic comprising said microcarrier bead and attached thereto or grown thereon at least one selected cell or tissue type; a method for making said therapeutic; and method a of treatment involving the use of said microcarrier bead or said therapeutic. 10 (Figure 1) 27 ...

SERUMFREE CELL CULTURES USEFUL FOR ANTIGENS PREPARATION AND PROCESS FOR PRODUCING FLAVIVIRUS/VIRUS ANTIGEN OR ARENAVIRUS/VIRUS ANTIGEN

RGD CONTAINING RECOMBINANT GELATIN

The invention concerns recombinant gelatins with unevenly distributed RGD motifs that are of particular use in several applications involving cell attachment such as in cell culture work and applications involving cell cultures of anchor dependent cells and also in a variety of medical applications.

PROCESS FOR EX VIVO EXPANSION OF STEM CELLS IN A BIOREACTOR

The present invention refers to a process of ex vivo expansion of stem cells, in a bioreactor, in particular hematopoietic stem/progenitor cells co-cultured with mesenchymal stem cells immobilized on microcarriers, for transplantation. The process comprises the steps of: a) forming a suspension of mesenchymal stem cells immobilized on microcarriers, b) inoculating in a bioreactor containing an expansion medium, hematopoietic cells co-cultured with mesenchymal stem cells immobilized on microcarriers c) expansion of hematopoietic cells. The process of the invention is capable of being implemented in a Kit.

METHOD FOR STEM CELL CULTURE AND CELLS DERIVED THEREFROM

There is described a method of promoting the attachment, survival and/or proliferation of a stem cell in culture, the method comprising culturing a stem cell on a positively-charged support surface. There are also provided a cell composition prepared according to the method of the invention.

METHOD FOR CONTROLLING BINDING OF CELLS TO A SUBSTRATE

The invention relates to a method for promoting the adhesion of cells to a substrate to which these cells usually have no or only low affinity, wherein the adhesion of the cells to the substrate is promoted by supplying the cells with the non-muscle myosin II inhibitor Blebbistatin so as to enable the cells to attach to surfaces to which they otherwise would not have sufficient affinity. Surprisingly, supplying the cells with the inhibitor enhances the capability of these cells to attach to surfaces to which they usually have no or only low affinity, for example, PTFE (Teflon®). The invention further concerns uses of the non- muscle myosin II inhibitor Blebbistatin and devices having at least one surface which is coated with cells that have no or only low affinity to said surface.

METHOD FOR PRODUCING A HETEROLOGOUS SECRETED PROTEIN FROM CHINESE HAMSTER OVARY CELLS GROWN ON MICROCARRIERS

The present invention relates to a method for producing a heterologous secreted protein from transformed CHO cells grown on microcarriers in a serum-free cell culture medium. This method avoids a production lag phase when switching from a serum-containing to a serum-free environment, a problem observed in the mammalian adherent cell culture production of secreted proteins.

Vacsularized tissue for transplantation

Tissue engineering holds enormous potential to replace or restore function to a wide range of tissues. However, the most successful applications have been limited to thin avascular tissues in which delivery of essential nutrients occurs primarily by diffusion. Pursuant to the present invention, a prevascularized, thick tissue construct is created having a network of capillaries with lumens capable of nutrient and origin delivery and forming anastamoses to host vasculature. A tissue transplantation strategy is comprised of (1) in vitro vascularization of a tisue construct, (2) transplantation of prevascularized tissue to wound bed of host where vessels of implantable tissue and host rapidly anastomose, and (3) host-directed remodeling and reorganization of the tissue and vascular network.

Disease model incorporation into an artificial immune system (AIS)

The present invention relates to methods for preparing an artificial immune system. The artificial immune system comprises a cell culture comprising a three-dimensional matrix comprising lymphoid tissue, a three-dimensional matrix comprising epithelial and/or endothelial cells, and diseased cells. The artificial immune system of the present invention can be used for in vitro testing of vaccines, adjuvants, immunotherapy candidates, cosmetics, drugs, biologics and other chemicals.

Method for allogeneic cell therapy

A method of manipulating allogeneic cells for use in allogeneic cell therapy providing a composition of highly activated allogeneic T-cells which are infused into immunocompetent cancer patients to elicit a novel anti-tumor immune mechanism, or Mirror Effect. In contrast to current allogeneic cell therapy protocols where T-cells in the graft mediate the beneficial graft vs. tumor (GVT) and detrimental graft vs. host (GVH) effects, the allogeneic cells of the present invention stimulate host T-cells to mediate the mirror of these effects. The mirror of the GVT effect is the host vs. tumor (HVT) effect. The mirror of the GVH effect is the host vs. graft (HVG) effect The anti-tumor HVT effect occurs in conjunction with a non-toxic HVG rejection effect. The highly activated allogeneic cells of the invention can be used to stimulate host immunity in a complete HLA mis-matched setting in a patient.

Biodegradable T-cell activation method

A biodegradable device for activating T-cells includes a biodegradable support and a binder attached to the biodegradable support, the binder having reactivity to one or more agents capable of binding to a T-cell surface antigen.

СПОСОБ ЭКСПАНСИИ КЛЕТОК, СПОСОБ ПОЛУЧЕНИЯ КОНДИЦИОННОЙ СРЕДЫ, ПОПУЛЯЦИЯ АДГЕЗИВНЫХ МЕЗЕНХИМАЛЬНЫХ СТРОМАЛЬНЫХ КЛЕТОК ПЛАЦЕНТЫ ИЛИ ЖИРОВОЙ ТКАНИ, ФАРМАЦЕВТИЧЕСКАЯ КОМПОЗИЦИЯ И ПРИМЕНЕНИЕ АДГЕЗИВНЫХ МЕЗЕНХИМАЛЬНЫХ СТРОМАЛЬНЫХ КЛЕТОК ПЛАЦЕНТЫ ИЛИ ЖИРОВОЙ ТКАНИ В ТРАНСПЛАНТАЦИИ

Изобретение относится к области биотехнологии, конкретно к получению мезенхимальных стромальных клеток плаценты или жировой ткани, и может быть использовано в медицине. Способ экспансии клеток включает культивирование адгезивных мезенхимальных стромальных клеток из плаценты или жировой ткани в трехмерных условиях культивирования, которые содействуют экспансии клеток. Изобретение позволяет получить адгезивные мезенхимальные стромальные клетки плаценты или жировой ткани, которые возможно использовать для поддержания гематопоэтических стволовых клеток, для иммуносупрессии, для получения кондиционных сред и в транспланталогии. 6 н. и 16 з.п. ф-лы, 22 ил., 3 табл.

СУСПЕНДИРОВАНИЕ И КЛАСТЕРИЗАЦИЯ ПЛЮРИПОТЕНТНЫХ КЛЕТОК ЧЕЛОВЕКА С ЦЕЛЬЮ ИХ ДИФФЕРЕНЦИРОВКИ В ПАНКРЕАТИЧЕСКИЕ ЭНДОКРИННЫЕ КЛЕТКИ

Изобретение относится к области биотехнологии, а именно к дифференцировке клеточного кластера плюрипотентных стволовых клеток в клетки дефинитивной энтодермы. Способ включает получение клеточного кластера плюрипотентных стволовых клеток из плюрипотентных стволовых клеток, выращенных на плоской прикрепленной культуре, сформированного из скопления клеток или не выделенного из суспензий отдельных клеток, причем плюрипотентные стволовые клетки представляют собой плюрипотентные стволовые клетки неэмбрионального происхождения, индуцибельные плюрипотентные клетки, репрограмированные плюрипотентные клетки, клетки, выделенные из соматических клеток взрослого человека, индуцированные плюрипотентные стволовые клетки, клетки, полученные из амниотической жидкости человека, человеческие партеноты или клетки из линий эмбриональных стволовых клеток человека H1, H7, H9 или SA002. Затем осуществляют перемещение клеточного кластера плюрипотентных стволовых клеток из плоской прикрепленной культуры в динамичную ...

СПОСОБЫ ЭКСПАНСИИ КЛЕТОК И ПРИМЕНЕНИЕ КЛЕТОК И КОНДИЦИОННЫХ СРЕД, ПОЛУЧЕННЫХ ТАКИМ ОБРАЗОМ, ДЛЯ ЛЕЧЕНИЯ

... 1. Способ экспансии клеток, причем способ включает культивирование адгезивных клеток из плаценты или жировой ткани в трехмерных условиях культивирования, которые содействуют экспансии клеток. ! 2. Способ по п.1, в котором трехмерная культура включает 3D-биореактор. ! 3. Способ п.1, в котором культивирование осуществляют, по меньшей мере, в течение 3 дней. ! 4. Способ по п.1, в котором указанное культивирование осуществляют до достижения, по меньшей мере, 60% конфлюэнтности. ! 5. Способ по п.1, где указанные адгезивные клетки содержат чип экспрессии позитивных маркеров, выбранный из группы, состоящей из CD73, CD90, CD29 и CD105. ! 6. Способ по п.1, где указанные адгезивные клетки содержат чип экспрессии негативных маркеров, выбранный из группы, состоящей из CD45, CD80, HLA-DR, CD11b, CD14, CD19, CD34 и CD79. ! 7. Способ по п.1, где указанные адгезивные клетки или среда характеризуются более высокой иммуносуппрессирующей активностью, чем активность адгезивных клеток плаценты или жировой ткани ...

Microfluidic model of the blood brain barrier

Improved culture method using integrin agonist

BIOLOGICAL ARTIFICIAL LIVER

WITH CELLS CARRIER COATED

PROCEDURE FOR HERSTELLLUNG REKOMBINANTER PROTEINS IN EUKARYONTI CELLS

VERFAHREN ZUR HERSTELLUNG VON FSME-VIRUS-ANTIGEN

THREE-DIMENSIONAL CULTIVATED FABRICS AND THEIR USE

NOT NATURAL ONE REKOMBINANTE GELATINEN WITH EXTENDED FUNCTIONALITY

PROCEDURE FOR THE PRODUCTION OF A HETEROLIED SEZERNIERTEN PROTEIN OUT ON MICRO CARRIERS GROWN CHO CELLS

A METHOD TO THE TRANSPLANT OF CELLS IN THE BRAIN AND YOUR THERAPEUTIC USE

Conditioned medium comprising Wnt proteins to promote repair of damaged tissue

Cell-coated supports

Biological artificial liver

CELL CULTURE MICROCARRIERS

BIO-REACTOR CELL CULTURE PROCESS

Co-culture lymphoid tissue equivalent (LTE) for an artificial immune system (AIS)

In vitro expansion of postpartum-derived cells using microcarriers

Compositions and methods for the growth and expansion of mammalian cells in culture are provided. In particular, methods for the growth and expansion of postpartum-derived cells ...

Method for culturing MDCK cells

The present invention pertains to: cloned MDCK cells which exhibit an expansion factor of 4.5-fold or more when being cultured using microcarriers; a method for culturing the MDCK cells; and a method for multiplying a virus by using the MDCK cell culturing method.

Suspension and clustering of human pluripotent stem cells for differentiation into pancreatic endocrine cells

The present invention provides methods of preparing aggregated pluripotent stem cell clusters for differentiation. Specifically, the invention discloses methods of differentiating pluripotent cells into beta cell, cardiac cell and neuronal cell lineages using suspension clustering. The methods involve preparing the aggregated cell clusters followed by differentiation of these clusters.

Pluripotent stem cell culture on micro-carriers

The present invention is directed to methods for the growth, expansion and differentiation of pluripotent stem cells on micro-carriers.

Culture substrate, method for manufacturing culture substrate, and culturing method and culturing device for stem cell

The present disclosure provides a culture substrate for culturing a stem cell, the culture substrate being provided with a surface section having soft regions extending side-by-side along a plurality of directions crossing each other; and a plurality of hard regions divided by the soft regions, wherein, in the surface section, the hard regions have acute sections protruding toward the soft regions, and the cell can be deformed into a shape that can be accommodated in the hard regions.

Suspension and clustering of human pluripotent cells for differentiation into pancreatic endocrine cells

The present invention provides methods of preparing aggregated pluripotent stem cell clusters for differentiation.

Suspension and clustering of human pluripotent stem cells for differentiation into pancreatic endocrine cells

The disclosure provides methods of preparing aggregated pluripotent stem cell clusters for differentiation to endoderm progenitor cells, pancreatic endocrine cells, mesoderm cells or ectoderm cells. Specifically, the disclosure provides methods of differentiating pluripotent cells into beta cell, cardiac cell and neuronal cell lineages using suspension clustering. The methods involve preparing the aggregated cell clusters followed by differentiation of these clusters.

MACROCARRIER

A macrocarrier for the propagation of biological cells is described. The macrocarrier comprises substrate particles that are coated with a thermoresponsive polymer, which is capable of providing the macrocarrier with a cell-receiving surface and responding to a change in temperature to release cells from the macrocarrier. At least 50% of the substrate particles have a particle size of at least 1 mm. A system for the propagation of biological cells and a process for the propagation of biological cells are also described.

CELL-CULTURE AND POLYMER CONSTRUCTS

Cells grown on a microcarrier are separated from the microcarrier by enzymatically digesting the microcarrier. More specifically, chondrocytes may be grown on dextran microcarrier beadlets and then the beadlets digested using dextranase to separate the chondrocytes from the carrier. Cells can also be grown on chitosan microcarriers to be used for implantation. In addition, cells can be grown on polysaccharide polymers to be used as implant devices. Various polymers serve as scaffolds for cells to be used for implantation. The polymers can be used for cell culture as well as for preparing scaffolds useful for tissue replacement such as cartilage tissue.

METHODS FOR CELL EXPANSION AND USES OF CELLS AND CONDITIONEDMEDIA PRODUCED THEREBY FOR THERAPY

A method of cell expansion is provided. The method comprising culturing adherent cells from placenta or adipose tissue under three-dimensional culturing conditions, which support cell expansion.

DIFFERENTIATION OF PLURIPOTENT STEM CELLS

The present invention is directed to methods to differentiate pluripotent stem cells. In particular, the present invention is directed to methods and compositions to differentiate pluripotent stem cells into cells expressing markers characteristic of the definitive endoderm lineage comprising culturing the pluripotent stem cells in medium comprising a sufficient amount of GDF-8 to cause the differentiation of the pluripotent stem cells into cells expressing markers characteristic of the definitive endoderm lineage.

METHODS FOR CULTURING UNDIFFERENTIATED CELLS USING SUSTAINED RELEASE COMPOSITIONS

Methods for culturing undifferentiated mammalian cells, such as stem and progenitor cells, are provided. The methods involve incubating the cell in the presence of a sustained release composition containing at least one growth factor, wherein the sustained release composition continuously releases the growth factor(s), and wherein the presence of the sustained level of growth factor maintains the cell in an undifferentiated state.

METHOD FOR CONTROLLING BINDING OF CELLS TO A SUBSTRATE

The invention relates to a method for promoting the adhesion of cells to a substrate to which these cells usually have no or only low affinity, wherein the adhesion of the cells to the substrate is promoted by supplying the cells with the non-muscle myosin II inhibitor Blebbistatin so as to enable the cells to attach to surfaces to which they otherwise would not have sufficient affinity. Surprisingly, supplying the cells with the inhibitor enhances the capability of these cells to attach to surfaces to which they usually have no or only low affinity, for example, PTFE (Teflon®). The invention further concerns uses of the non- muscle myosin II inhibitor Blebbistatin and devices having at least one surface which is coated with cells that have no or only low affinity to said surface.

PROCESS FOR EX-VIVO EXPANSION OF HEMATOPOIETIC STEM CELLS IN A BIOREACTOR

The present invention refers to a process of ex vivo expansion of stem cells, in a bioreactor, in particular hematopoietic stem/progenitor cells co-cultured with mesenchymal stem cells immobilized on microcarriers, for transplantation. The process comprises the steps of: a) forming a suspension of mesenchymal stem cells immobilized on microcarriers, b) inoculating in a bioreactor containing an expansion medium, hematopoietic cells co-cultured with mesenchymal stem cells immobilized on microcarriers c) expansion of hematopoietic cells. The process of the invention is capable of being implemented in a Kit.

CO-CULTURE LYMPHOID TISSUE EQUIVALENT (LTE) FOR AN ARTIFICIAL IMMUNE SYSTEM (AIS)

The present invention relates to methods for preparing an artificial immune system. The artificial immune system comprises a cell culture comprising T cells, B cells and antigen-primed dendritic cells. The artificial immune system of the present invention can be used for in vitro testing of vaccines, adjuvants, immunotherapy candidates, cosmetics, drugs, biologies and other chemicals.

СПОСОБ ПОЛУЧЕНИЯ ГЕТЕРОЛОГИЧНОГО СЕКРЕТИРУЕМОГО БЕЛКА ИЗ КЛЕТОК ЯИЧНИКА КИТАЙСКОГО ХОМЯЧКА, ВЫРАЩИВАЕМЫХ НА МИКРОНОСИТЕЛЯХ

... 1. Способ получения гетерологичного секретируемого белка из клеток яичника китайского хомячка (CHO), выращиваемых на микроносителях, предусматривающий инокуляцию клеточного биореактора, содержащего микроносители в качестве твердой подложки для роста прикрепленных клеток и содержащую сыворотку среду для культуры клеток, инокулятом хозяйских клеток CHO, трансформированных для экспрессии гетерологичного секретируемого белка, где инокулят получен путем выращивания трансформированных хозяйских клеток CHO в содержащей сыворотку среде для культуры клеток; и культивирование инокулированных хозяйских клеток CHO в культуре прикрепленных клеток без перфузии среды для культуры клеток в течение периода приблизительно от одного до трех дней до инициации фидинга (подпитки); введение усовершенствования, подразумевающего инициацию фидинга бессывороточной среды для культуры клеток, дополненной инсулином, цитратом железа (III), селеном и микроэлементами, в клеточный биореактор путем непрерывной перфузии; ...

METHOD FOR PREPARING HETEROLOGOUS SECRETED PROTEIN FROM CHINESE HAMSTER OVARY CELLS, GROWN ON MICROCARRIERS

Method of producing lymphocytes

A method of producing lymphocytes characterized by comprising the step of performing enlarged culture in the presence of (a) fibronectin, its fragment or a mixture thereof, (b) CD3 ligand and (c) CD28 ligand.

Methods for treating radiation or chemical injury

Reposiçao de pele viva e processos para a produçao da mesma e de uso de microesferas revestidas de células

SCALE UP OF CELL CULTURES

The invention relates to a method and device for cultivating attachment-dependent cells in the presence of microcarriers in a flexible cultivation container. In particular, the invention concerns a cultivation method using a flexible, scalable, disposable, substantially gas-permeable and non-liquid-permeable container for expansion of adherent cells on microcarriers. According to the invention, cultivation of the cells is accomplished under orbital shaking at 8 rpm or more than 8 rpm.

CELL CULTURE SYSTEM FOR BIOREACTOR SCALE-UP OF CELLS

The present invention relates to growing stem cells, for e.g., MSCs, in large - scale under GMP-compliance, using media and reagents that satisfy GMP requirements, while maintaining stemness, for effective downstream therapeutic use, which include but are not limited to, stem cell therapy, production of products, such as beneficial factors, recombinant proteins, etc. obtained from such stem cells.

THREE-DIMENSIONAL PERIPHERAL LYMPHOID ORGAN CELL CULTURES

The present invention relates to a method of culturing peripheral lymphoid organ cells on a three-dimensional scaffolding which is covered or surrounded with culture medium, under conditions effective to generate and maintain mature and functional lymphoid cells, where the three-dimensional scaffolding allows cells in the culture medium to have cell to cell contact in three dimensions. The present invention also provides methods of screening for vaccine candidates for efficacy in eliciting an immune response, identifying genes or proteins which are related to peripheral lymphoid organ cell formation or function, screening for drugs effecting peripheral lymphoid organ cell maturation, treating a patient for a disease condition using antigen-specific lymphoid cells, and effecting gene expression of peripheral lymphoid organ cells.

PEPTIDE-MODIFIED MICROCARRIERS FOR CELL CULTURE

A cell culture article including a microcarrier having a peptide-modified polymer surface of the formula (I) where AAj represents at least one covalently bonded peptide, j is an integer of from 5 to 50, m, n, o, Sur, X, R, R', and the mer ratio (m-o:n:o), including salts thereof, are as defined herein. Also disclosed are methods for making and using the cell culture article, as defined herein.

BIOSTABILITY OF POLYMERIC STRUCTURES

Selon l'invention, un matériau polymérique biocompatible est préparé par formation d'une structure réticulée tridimensionnel d'un matériau polymérique biocompatible, tel qu'un polyuréthane de polyéther ou de polycarbonate, et par extraction par solvant du matériau à l'aide d'un solvant de gonflement, tel que le MEK qui peut gonfler le matériau jusqu'à 150 %. Le matériau polymérique gonflé au solvant est ensuite dégonflé avec un produit non solvant, comme l'eau qui est miscible avec le solvant d'extraction. Le procédé donne des matériaux polymériques non lixiviables et possédant par conséquent des propriétés qui les rendent appropriés pour des implantations.

Manufacturing process for polysaccharide beads

The invention discloses a method of manufacturing polysaccharide beads, comprising the steps of: i) providing a water phase comprising an aqueous solution of a polysaccharide; ii) providing an oil phase comprising at least one water-immiscible organic solvent and at least one oil-soluble emulsifier; iii) emulsifying the water phase in the oil phase to form a water-in-oil (w/o) emulsion; and iv) inducing solidification of the water phase in the w/o emulsion, wherein the organic solvent is an aliphatic or alicyclic ketone or ether.

Peptide-modified microcarriers for cell culture

A cell culture article including a microcarrier having a peptide-modified polymer surface of the formula (I) where AAj represents at least one covalently bonded peptide, j is an integer of from 5 to 50, m, n, o, Sur, X, R, R′, and the mer ratio (m-o:n:o), including salts thereof, are as defined herein. Also disclosed are methods for making and using the cell culture article, as defined herein.

Differentiation of Pluripotent Stem Cells

The present invention is directed to methods to differentiate pluripotent stem cells. In particular, the present invention is directed to methods and compositions to differentiate pluripotent stem cells into cells expressing markers characteristic of the definitive endoderm lineage comprising culturing the pluripotent stem cells in medium comprising a sufficient amount of GDF-8 to cause the differentiation of the pluripotent stem cells into cells expressing markers characteristic of the definitive endoderm lineage.

Macroporous Microcarrier Specific to Liver Cell, Preparation Method and Use Thereof

The present invention provides a macroporous microcarrier specific to hepatocytes using silk fibroin and galactosylated chitosan as main raw material, a preparation method thereof, and application for hepatocyte culture under the culture condition of microgravity rotation. The macroporous microcarrier s a sphere prepared from silk fibroin and galactosylated chitosan under the effect of crosslinker, wherein based on the total weight of the sphere, the content of silk fibroin is 50-80 wt % and the content of galactosylated chitosan is 15-40 wt %. The diameter of the microcarrier is 200-500 μm, and the aperture of the microcarrier is 40-80 μm. Compared with normal solid scaffold material, the microcarrier provided by the present invention has larger surface area/volume ratio and, a sinus gap structure extremely similar with in-vivo liver sinus structure, therefore it is more conducive to adhering of the hepatocytes on the scaffold material, contacting between cells, transporting oxygen and nutrient components and excreting metabolic products.

Artificial immune system: methods of use

The present invention relates to methods of constructing an integrated artificial immune system that comprises appropriate in vitro cellular and tissue constructs or their equivalents to mimic the normal tissues that interact with vaccines in mammals. The artificial immune system can be used to test the efficacy of vaccine candidates in vitro and thus, is useful to accelerate vaccine development and testing drug and chemical interaction with the immune system.

MICROCARRIER PERFUSION CULTURING METHODS AND USES THEREOF

Provided herein are methods of culturing a mammalian cell and various methods that utilize these culturing methods. 1. A method of culturing a mammalian cell , the method comprising:providing a shake flask containing a mammalian cell disposed in a first liquid culture medium, wherein the first liquid culture medium occupies about 20% to about 30% of the volume of the shake flask and contains a plurality of microcarriers at a concentration of about 1.0 g/L to about 15.0 g/L;incubating the shake flask for a period of time at about 32° C. to about 39° C. and with a rotary agitation of about 85 revolutions per minute (RPM) to about 125 RPM; andafter about the first 48 to 96 hours of the period of time, continuously or periodically removing a first volume of the first liquid culture medium and adding to the first liquid culture medium a second volume of a second liquid culture medium, wherein the first and second volumes are about equal.2. The method of claim 1 , wherein the first volume of the first liquid culture medium is substantially free of the microcarriers.3. The method of claim 1 , wherein the first liquid culture medium occupies about 25% to about 30% of the volume of the shake flask.4. The method of claim 1 , wherein at the beginning of the period of time claim 1 , the first liquid culture medium contains 0.1×10cells/mL to 0.5×10cells/mL.5. The method of claim 1 , wherein the mammalian cell is a Chinese hamster ovary (CHO) cell.6. The method of claim 5 , wherein the CHO cell contains a nucleic acid encoding a recombinant protein.7. The method of claim 5 , wherein the recombinant protein is an immunoglobulin claim 5 , an enzyme claim 5 , a growth factor claim 5 , a protein fragment claim 5 , or an engineered protein.8. The method of claim 1 , wherein the removing of the first volume of the first liquid culture medium and the adding of the second volume of the second liquid culture medium is performed simultaneously.9. The method of claim 1 , wherein the ...

CONTROL OF CELL GROWTH AND AGGREGATE SIZE IN BIOREACTORS

Methods of repeated aggregate dissociation and reformation of pluripotent stem cells (PSCs) within the same bioreactor until a desired final cell number is achieved. A preferred step-wise process for controlled growth of PSCs and aggregate size using periodic dissociation with a dissociation medium which contains either proteolytic enzymes or chemical reagents, mechanical agitation, or a combination of these methods. 1. A method of controlling the growth of cells and aggregates thereof , comprising:a. seeding a bioreactor with anchorage-dependent cells;b. operating the bioreactor for an initial period of time such that the cells form cell aggregates that will continue to grow in size and thus also increase the total number of cells in the bioreactor;c. dissociating the cell aggregates within the same bioreactor; andd. repeating the steps of operating and dissociating within the same bioreactor until a desired number of cells has been reached or the capacity of the bioreactor is fully utilized.2. The method of claim 1 , wherein the cells are selected from the group consisting of pluripotent stem cells (PSCs) claim 1 , mesenchymal stem cells (MSCs) claim 1 , human primary cells claim 1 , or any other anchorage-dependent cells that require aggregate formation for growth.3. The method of where dissociating the cell aggregates can be accomplished using either a dissociation medium containing proteolytic enzymes or chemical reagents claim 1 , mechanical agitation claim 1 , or a combination of these methods.4. The method of where the timing of dissociation is dictated by cell aggregates reaching a predetermined threshold size claim 1 , or range of sizes.5. A method of controlling the growth of cells and aggregates thereof claim 1 , comprising:a. seeding a bioreactor with anchorage-dependent cells as suspended single cells, along with microcarriers;b. operating the bioreactor for an initial period of time such that the cells first attach to the surface of microcarriers ...

SYSTEM FOR PRODUCING CULTIVATED MEATS, TISSUES AND ASSOCIATED PRODUCTS FROM CELLS

A cell/tissue culture system comprising at least one bioreactor configured to hold at least one type of cell to cultivate tissue, a dialysis unit comprising a dialysis membrane, a fresh medium unit, and a waste removal unit configured to remove metabolic waste from dialysate, wherein the metabolic waste comprises ammonia and lactate, and wherein the waste removal unit comprises biocatalysts or enzymes configured to breakdown lactate and generate carbon sources that promote cell growth. 1. A system for in vitro meat production comprising:a bioreactor configured to hold at least one type of cells to form tissue;a dialysis unit comprising a dialysis membrane configured to replenish nutrients to the cells and remove metabolic waste from the bioreactor to dialysate;a fresh medium unit connected to the dialysis unit and configured to supply the dialysate to the dialysis unit; anda waste removal unit connected to the fresh medium unit and configured to remove the metabolic waste from the dialysate;wherein the dialysate comprises nutrients for the cells,wherein the metabolic waste comprises ammonia and lactate,wherein the waste removal unit comprises a first biocatalyst configured to breakdown ammonia,wherein the waste removal unit comprises a second biocatalyst configured to breakdown lactate,wherein each of the bioreactors, the dialysis unit, the fresh medium unit and the water removal unit are detachably connected to the system.2. The system of claim 1 , wherein the second biocatalyst is an enzyme configured to break down lactate.3. The system of claim 1 , wherein the waste removal unit is further configured to generate fresh carbon sources that can promote cell growth in the bioreactor.4. The system of claim 3 , wherein the fresh carbon source is pyruvate.5. The system of claim 1 , wherein the first biocatalyst is nitrifying bacteria.6. The system of claim 1 , wherein the dialysis membrane is at least one 500 Da molecular weight cut-off (MWCO) membrane claim 1 , wherein ...

ORGANOID ARRAYS

The invention provides methods for producing arrays of organoids, the arrays thereof and uses of such arrays. 1. A method for making an array of organoids , comprising:i. seeding stem cells on a surface,ii. culturing the stem cells of step i) in situ to allow their aggregation into multicellular stem cell containing aggregates,iii. culturing the multicellular stem cell containing aggregates of ii) in situ in conditions suitable for organoid development,wherein the array of organoids is within a single focal plane and the surface may be overlaid with a regular tessellation, such that the organoids are uniquely positioned within adjacent tiles of the tessellation; and wherein the surface comprises a biofunctional hydrogel.2. The method of claim 1 , wherein non-stem cells are seeded in combination with the stem cells seeded in step i claim 1 , and/or additionally comprising overlaying the multicellular stem cell containing aggregates with an overlay claim 1 , wherein the overlay comprises a gel or viscous solution claim 1 ,preferably wherein the overlay comprises a cell compatible material that supports organoid development and maintenance,more preferably wherein the cell compatible material is a hydrogel, preferably wherein the viscous solution is a dilute hydrogel,most preferably wherein the surface comprises a hydrogel that has a stiffness between 150 Pa and 50 kPa.3. The method of claim 2 , wherein the surface hydrogel and/or overlay hydrogel comprises naturally derived biomaterials claim 2 , i. polysaccharides, gelatinous proteins, ECM components comprising: agarose; alginate; chitosan; dextran; gelatin; lam inins; collagens; hyaluronan; fibrin, functional variants thereof, and mixtures thereof; or', 'ii. a gel derived from natural ECM, preferably Matrigel, Myogel or Cartigel., 'preferably wherein the naturally derived biomaterials are selected from the group comprising4. The method of claim 2 , wherein the surface hydrogel and/or overlay hydrogel is a crosslinked ...

Cell culture substrate, culture vessel, method for producing cell culture vessel, method for acquiring cells and method for culturing cells

A cell culture substrate includes: a first layer that includes a first gel in which gold nanoparticles dispersed; and a second layer that includes a second gel in which the gold nanoparticles are not present or are present in a lower concentration in comparison with the first layer.

METHODS FOR CELL EXPANSION AND USES OF CELLS AND CONDITIONED MEDIA PRODUCED THEREBY FOR THERAPY

A method of cell expansion is provided. The method comprising culturing adherent cells from placenta or adipose tissue under three-dimensional culturing conditions, which support cell expansion. 141-. (canceled)42. A method for preparing a pharmaceutical composition , comprisingintroducing adherent stromal cells (ASC) into a growth medium containing a plurality of 3D carriers, wherein the 3D carriers comprise an adherent material selected from the group consisting of a polyester, a polyalkylene, a polyfluorochloroethylene, a polyvinyl chloride, a polystyrene, and a polysulfone,incubating the growth medium containing the 3D carriers in a bioreactor, andremoving the ASC from the 3D carriers,thereby preparing a pharmaceutical composition.43. The method of claim 42 , wherein the 3D carriers are submerged in the growth medium.44. The method of claim 42 , wherein the 3D carriers are suspended in the growth medium.45. The method of claim 42 , wherein the 3D carriers are microcarriers.46. The method of claim 42 , wherein the 3D carriers comprise a non-woven fibrous matrix.47. The method of claim 42 , wherein the 3D carriers are packed in the bioreactor.48. The method of claim 42 , whereby the ASC are expanded on the 3D carriers.49. The method of claim 42 , wherein the ASC adhere to the 3D carriers.50. The method of claim 42 , wherein the ASC are in the form of a cell suspension in the pharmaceutical composition.51. The method of claim 42 , wherein the 3D carriers comprise polystyrene.52. The method of claim 42 , wherein the 3D carriers substantially consist of polystyrene.53. The method of claim 42 , wherein the surface of the 3D carriers comprises the adherent material.54. The method of claim 53 , wherein the adherent material is polystyrene.55. The method of claim 42 , wherein the step of removing takes place in the bioreactor.56. The method of claim 42 , wherein the ASC are derived from bone marrow.57. The method of claim 42 , wherein the ASC are derived from cord blood. ...

Adipose tissue-derived stem cells for veterinary use

The invention provides for compositions and methods for making and using adipose-derived stem cells for treating non-human mammals for various medical conditions.

METHOD FOR STEM CELL CULTURE AND CELLS DERIVED THEREFROM

There is described a method of promoting the attachment, survival and/or proliferation of a stem cell in culture, the method comprising culturing a stem cell on a positively-charged support surface. There are also provided a cell composition prepared according to the method of the invention. 1. A cell culture system comprising substantially undifferentiated stem cells cultured on a positively-charged support surface.2. The cell culture system of claim 1 , wherein the support surface comprises a positively-charged molecule bound thereto and wherein the positively charged molecule is tri-methylamine.3. The cell culture system of claim 1 , wherein the support surface is selected from the group consisting of a tissue culture plate claim 1 , a microscope slide claim 1 , a multi-well plate claim 1 , a flask claim 1 , a bottle claim 1 , a bioreactor claim 1 , a two- or three-dimensional scaffold claim 1 , a tube claim 1 , a suture claim 1 , a membrane claim 1 , a film claim 1 , a microcarrier bead claim 1 , a tissue and an organ.4. The cell culture system of claim 3 , wherein the support surface is the surface of a microcarrier bead.5. The cell culture system of claim 1 , further comprising an extracellular matrix component.6. The cell culture system of claim 5 , wherein the extracellular matrix component is selected from the group consisting of elastin claim 5 , fibronectin claim 5 , vitronectin claim 5 , tenascin claim 5 , laminin claim 5 , entactin claim 5 , aggrecan claim 5 , decorin claim 5 , collagen I claim 5 , collagen III claim 5 , collagen IV claim 5 , and collagen VI claim 5 , biologically active fragments or variants of said proteins claim 5 , or combinations thereof.7. The cell culture system of claim 1 , wherein the substantially undifferentiated stem cells are passaged and maintained as single-cell culture.8. The cell culture system of claim 1 , wherein the substantially undifferentiated stem cells are human stem cells.9. The cell culture system of claim 1 , ...

Methods for Preparing Therapeutically Active Cells Using Microfluidics

The present invention is directed to the use of microfluidics in the preparation of cells and compositions for therapeutic uses.

METHODS AND KITS FOR DIRECTING CELL ATTACHMENT AND SPREADING

The present disclosure provides methods, and related kits, for directing cell attachment and spreading on a substrate and inducing isotropic spreading of cells; provides methods, and related kits, for cell sorting; and further provides methods, and related kits, for guided induction of stem cell differentiation. 1. A method of directing cell attachment and spreading on a substrate , comprising:culturing a cell on a curved substrate in the presence of cell culture media, wherein attachment and spreading increases as the curvature of the substrate decreases.2. The method of claim 1 , wherein the curved substrate is selected from the group consisting of a convex substrate claim 1 , a concave substrate claim 1 , a spherical substrate claim 1 , an oval substrate claim 1 , an elliptical substrate claim 1 , and combinations thereof.3. The method of claim 2 , wherein the curved substrate is a spherical substrate claim 2 , wherein the attachment and spreading increases as the diameter of the spherical substrate increases.4. The method of claim 3 , wherein the spherical substrate comprises a diameter of between about 500 μm and about 6 mm claim 3 , wherein attachment and spreading is influenced in that both attachment and spreading increases as the diameter of the spherical substrate increases from about 500 μm to about 6 mm.5. The method of claim 1 , wherein the curved substrate comprises a coating selected from the group consisting of a cell adhesive claim 1 , a cell adhesion-promoter claim 1 , a cell repellent claim 1 , or a combination thereof6. A method of inducing isotropic spreading of cells claim 1 , comprising:culturing a cell on a curved substrate, the curved substrate immobilized on a material layer, wherein the cell isotropically spreads over the curved substrate and onto the material layer.7. The method of claim 6 , wherein the curved substrate is a spherical substrate.8. The method of claim 7 , wherein the spherical substrate comprises a diameter of between ...

METHODS AND KITS FOR GUIDED STEM CELL DIFFERENTIATION

The present disclosure provides methods, and related kits, for directing cell attachment and spreading on a substrate and inducing isotropic spreading of cells; provides methods, and related kits, for cell sorting; and further provides methods, and related kits, for guided induction of stem cell differentiation. 1. A method for guided induction of stem cell differentiation , comprising:culturing a stem cell on a curved substrate in the presence of cell culture media.2. The method of claim 1 , wherein the curved substrate comprises a coating selected from the group consisting of a cell adhesive claim 1 , a cell adhesion-promotor claim 1 , a cell repellent claim 1 , or a combination thereof.3. The method of claim 1 , wherein the curved substrate is selected from the group consisting of a convex substrate claim 1 , a concave substrate claim 1 , a spherical substrate claim 1 , an oval substrate claim 1 , an elliptical substrate claim 1 , and combinations thereof.4. The method of claim 1 , wherein the curved substrate is a spherical substrate.5. The method of claim 4 , wherein the spherical substrate comprises a diameter of between about 500 μm and about 4 mm.6. The method of claim 4 , wherein the spherical substrate comprises a diameter of between about 500 μm and about 2 mm.7. The method of claim 4 , wherein the spherical substrate comprises a diameter of between about 4 mm and about 6 mm.8. The method of claim 4 , wherein the spherical substrate comprises a diameter of between about 500 μm and about 6 mm.9. The method of claim 1 , wherein the stem cell is a mesenchymal stem cell.10. The method of claim 9 , wherein the cell culture media comprises osteocyte differentiation induction media claim 9 , whereby the stem cell differentiates into an osteocyte.11. The method of claim 9 , wherein the stem cell differentiates into an adipocyte.12. A kit for guided induction of stem cell differentiation claim 9 , comprising:a first reagent, wherein the first reagent comprises a ...

Methods, Compositions, and Systems for Activation and Expansion of Cells

The disclosure provides for compositions, systems, and methods of cell expansion, stimulation and/or differentiation. The disclosure further provides for a mesh substrate and associated methods capable of stimulating cell expansion, for example, T cell or stem cell expansion. In another aspect, the disclosure provides for an electrospun mesh substrate and methods of using thereof comprising a silicone rubber composition, for example, polydimethylsiloxane, PLC, or combinations thereof. 1. A method of improving cell expansion comprising culturing cells on a mesh substrate comprising polydimethylsiloxane and polycaprolactone in a ratio , wherein said mesh comprises fibers with a diameter of about 100 nm to about 10 μm and a pore size of about 0.5 μm to about 100 μm , wherein said cells are T-cells.2. The method of claim 1 , wherein said mesh substrate comprises fibers with a diameter selected from the group consisting of about 100 nm to about 1000 nm claim 1 , about 100 nm to about 2000 nm claim 1 , about 500 nm to about 5000 nm claim 1 , and about 1000 nm to about 5000 nm.3. The method of claim 1 , wherein said mesh substrate comprises fibers with a pore size selected from the group consisting of about 1 μm to about 100 μm claim 1 , about 1 μm to about 50 μm claim 1 , about 1 μm to about 10 μm claim 1 , and about 1 μm to about 5 μm.4. The method of claim 1 , wherein said cells arei. isolated from an individual,ii. expanded in vitro, andiii. transfused back to an individual in need thereof.5. The method of claim 4 , wherein the material comprising the fibers has a bulk rigidity that is greater than a local rigidity as indicated by atomic force microscopy indentation method.6. The method of claim 5 , wherein the material comprising the fibers has a bulk rigidity that is at least twice as high as a local rigidity as indicated by atomic force microscopy indentation method.7. The method of claim 6 , where in the bulk elastic modulus of the fiber is less than 10 MPa.8. The ...

IN VITRO EXPANSION OF ERYTHROID CELLS

The present invention relates to a method for in vitro expansion of erythroid cells. The method includes subjecting erythroid cells to 3-dimensional packed cell culture using a porous structure. The use of the composition according to the present invention enables in vitro expansion of erythroid cells in the most efficient manner. 1. A method for in vitro expansion of erythroid cells comprising subjecting erythroid cells to 3-dimensional packed cell culture using a porous structure.2. The method according to claim 1 , wherein the porous structure has a size distribution of 30 to 500 μm.3. The method according to claim 1 , wherein the culture is performed in a medium to which shear stress is applied by a continuous flow.4. The method according to claim 3 , wherein the flow is created by stirring.5. The method according to claim 4 , wherein the flow is created by stirring at 1 to 50 rpm.6. The method according to claim 3 , wherein the culture is performed in a filter in the medium to prevent erythrocytes from escaping from the porous structure during the continuous flow.7. The method according to claim 6 , wherein the filter has a mesh size of 1 to 8 μm.8. The method according to claim 1 , wherein the erythroid cells are cells that enter the terminal maturation stage.9. The method according to claim 1 , wherein the 3D-cultured erythroid cells express at least one adhesion-related gene selected from deleted in liver cancer 1 (DLC 1) claim 1 , intercellular adhesion molecule-4 (ICAM-4) claim 1 , and very late antigen-4 (VLA-4).10. The method according to claim 1 , wherein the erythroid cells are mixed and co-cultured with cells selected from the group consisting of mesenchymal stem cells claim 1 , endothelial cells claim 1 , monocytes claim 1 , macrophages claim 1 , and histiocytes during the 3-dimensional packed cell culture.11. The method according to claim 10 , wherein the erythroid cells are mixed and co-cultured with mesenchymal stem cells claim 10 , endothelial ...

BIOPHOSPHONATE COMPOUNDS AND GAMMA DELTA T CELL-MEDIATED THERAPY FOR TREATING EPSTEIN-BARR VIRUS-ASSOCIATED DISORDERS

Aminobisphosphonate pamidronate (PAM) can control Epstein-Barr virus (EBV) associated disorders in humanized mice through a Vγ9Vδ2-T-cell dependent mechanism. This suggests a strong potential for a therapeutic approach using PAM to boost human Vγ9Vδ2-T-cell immunity against EBV associated disorders, such as the lymphoproliferative disease (LPD), posttransplant lymphoproliferative disorder (PLPD), Hodgkin's disease, Burkitt's lymphoma, and nasopharyngeal carcinoma (NPC). 1. A method of treating a subject having an Epstein-Barr virus (EBV)-associated disorder , comprising administering to the subject an effective amount of aminobisphosphonate pamidronate (PAM).2. The method of claim 1 , wherein the EBV-associated disorder is selected from lymphoproliferative disease (LPD) claim 1 , posttransplant lymphoproliferative disorder (PLPD) claim 1 , Hodgkin's disease claim 1 , Burkitt's lymphoma claim 1 , and nasopharyngeal carcinoma (NPC).3. The method of claim 1 , wherein the subject is a human.4. The method of claim 1 , wherein PAM is administered with a pharmaceutically acceptable carrier.5. A method of treating a subject having an Epstein-Barr virus (EBV)-associated disorder claim 1 , comprising administering to the subject an effective amount of aminobisphosphonate pamidronate (pam)-expanded Vγ9Vδ2-T cells.6. The method of claim 5 , wherein the subject is a human.7. The method of claim 5 , wherein the PAM-expanded Vγ9Vδ2-T cells are administered with a pharmaceutically acceptable carrier.8. A pharmaceutical composition comprising aminobisphosphonate pamidronate (PAM) and a pharmaceutically acceptable carrier.9. The pharmaceutical composition of claim 8 , further comprising one or more pharmaceutically acceptable excipients claim 8 , additives claim 8 , or adjuvants.10. A pharmaceutical composition comprising aminobisphosphonate pamidronate (PAM)-expanded Vγ9Vδ2-T cells and a pharmaceutically acceptable carrier.11. The pharmaceutical composition of claim 10 , further ...

Decellularized Tissue as a Microcarrier for Cell Culture and Expansion

A microcarrier for cell culture and expansion is provided. The microcarrier includes decellularized mammalian tissue. Further, the microcarrier has an average particle size ranging from about 10 micrometers to about 600 micrometers. A method of forming a decellularized mammalian tissue microcarrier for cell culture and expansion is also provided, along with a method for treating a mammalian tissue defect via a decellularized mammalian tissue microcarrier on which cells from the same tissue type as the decellularized mammalian tissue are expanded. 1. A microcarrier for cell culture and expansion comprising decellularized mammalian tissue , wherein the microcarrier has an average particle size ranging from about 10 micrometers to about 600 micrometers.2. The microcarrier of claim 1 , wherein the decellularized mammalian tissue originates from a human donor claim 1 , a specific human patient claim 1 , or a non-human mammal.3. The microcarrier of claim 1 , wherein the decellularized mammalian tissue originates from embryonic tissue claim 1 , neonatal tissue claim 1 , natal tissue claim 1 , juvenile tissue claim 1 , or adult tissue.4. The microcarrier of claim 1 , wherein the decellularized mammalian tissue originates from articular cartilage tissue claim 1 , bone tissue claim 1 , heart tissue claim 1 , liver tissue claim 1 , skin tissue claim 1 , or gall bladder tissue.5. The microcarrier of claim 1 , wherein the microcarrier is compatible with cells harvested from mammalian tissue of a same type as the decellularized mammalian tissue.6. The microcarrier of claim 1 , wherein the decellularized mammalian tissue is micronized.7. The microcarrier of claim 1 , wherein the decellularized mammalian tissue is digested and functionalized.8. The microcarrier of claim 7 , wherein the decellularized mammalian tissue is cross-linked.9. The microcarrier of claim 1 , wherein the microcarrier has a honeycomb microstructure.10. The microcarrier of claim 1 , wherein the microcarrier ...

ADDITION OF NUCLEASES DIRECTLY TO CELL CULTURE TO FACILITATE DIGESTION AND CLEARANCE OF HOST CELL NUCLEIC ACIDS

The present invention provides an efficient process for culturing viruses in the presence of an endonuclease and for producing vaccines, typically from live attenuated viruses, under conditions to reduce the presence of host cell DNA and eliminate the need for a post-harvest DNA digestion step. 1. A method for production of a flavivirus , comprising the steps of:a) infecting a culture of cells in a cell culture medium with the flavivirus; andb) incubating the cells in cell culture medium containing endonuclease for up to 15 days, wherein the endonuclease is added to the cell culture medium at any time from initial infection of the cells to prior to harvesting the cells and wherein the cells are adherent to a microcarrier or a plastic surface.2. The method of claim 1 , wherein the endonuclease is added at the time of initial infection.3. The method of claim 1 , wherein the endonuclease is added from 0.5 to 15 days prior to harvesting the cells.4. The method of claim 3 , wherein the endonuclease is added 7 claim 3 , 8 claim 3 , 9 claim 3 , 10 claim 3 , 11 claim 3 , 12 claim 3 , 13 claim 3 , or 15 days prior to harvesting the cells.5. The method of claim 1 , wherein the endonuclease may have DNA specificity claim 1 , RNA specificity claim 1 , or both.6. The method of claim 5 , wherein the endonuclease is a broad spectrum endonuclease derived from the bacterium Serratia marcescens.7. The method of claim 1 , wherein the cell culture medium comprises endonuclease at an initial concentration of 2.5 to 80 U per mL of medium.8. The method of claim 7 , wherein the cell culture medium comprises endonuclease at an initial concentration of 2.5 to 20 U per mL.9. The method of claim 8 , wherein the cell culture medium comprises endonuclease at an initial concentration of 5 U per mL.10. The method of claim 1 , wherein the culture of cells are mammalian cells.11. The method of claim 10 , wherein the mammalian cells are adherent to a microcarrier.12. The method of claim 11 , wherein ...

SYSTEMS AND METHODS FOR CULTURING EPITHELIAL CELLS

The present invention features assays for co-culturing primary cells while maintaining key biological activities specific to the primary cells. The invention is based, at least in part, on the discovery that compositions and methods for primary cells in a high-throughput co-culture platform, image analysis for distinguishing cells in co-cultures and assays that are suitable for screening of agents in epithelial cells, such as hepatocytes. 132.-. (canceled)33. A method for assessing an agent that alters hepatocyte biological activity , the method comprising:contacting with an agent a hepatocyte present in a co-culture for high throughput analysis of primary hepatocytes; wherein the co-culture comprises:a layer of feeder cells disposed without aggregation in a well of a multi-well plate comprising at least 96 wells;a layer of primary hepatocytes disposed on the feeder cells at a concentration that prevents contact inhibition of the hepatocytes; andan amount of culture medium that supports the hepatocytes and maintains at least one hepatocyte biological activity, wherein the amount of culture medium is optimized to balance oxygen transport and nutrient supply in the co-culture; andassaying for an alteration in hepatocyte biological activity relative to the activity of a control hepatocyte not exposed to the agent, wherein detection of the alteration identifies the agent as altering hepatocyte biological activity.34. The method of claim 33 , wherein the hepatocyte biological activity is proliferation claim 33 , viability claim 33 , differentiation claim 33 , toxicity claim 33 , or cell death.35. The method of claim 33 , wherein the method further comprises measuring albumin output as a surrogate marker for protein synthesis; measuring urea generation as a surrogate marker for amino acid metabolism function; and/or measuring cytochrome P450 activity as a surrogate marker for detoxification.36. (canceled)37. The method of claim 33 , wherein the agent comprises one or more ...

Process for Ex Vivo Expansion of Stem Cells in a Bioreactor

The present invention refers to a process of ex vivo expansion of stem cells, in a bioreactor, in particular hematopoietic stem/progenitor cells co-cultured with mesenchymal stem cells immobilized on microcarriers, for transplantation. The process comprises the steps of: a) forming a suspension of mesenchymal stem cells immobilized on microcarriers, b) inoculating in a bioreactor containing an expansion medium, hematopoietic cells co-cultured with mesenchymal stem cells immobilized on microcarriers c) expansion of hematopoietic cells. The process of the invention is capable of being implemented in a Kit. 1. Process for rapid expansion of number of hematopoietic stem and progenitor cells , characterized in that it comprises simultaneously:a) providing a bioreactor system under stirred dynamic conditions;b) providing a co-culture of hematopoietic stem cells with mesenchymal stem cells on inert supports, in the presence of growth factors in serum free medium, constituting a cell culture;c) maintaining the cell culture in said bioreactor under dynamic conditions such that the expansion of hematopoietic stem cells is at least 5-fold in less than 20 days.2. Process according to characterized in that it comprises mesenchymal stem cells isolated from bone marrow or umbilical cord blood claim 1 , or from umbilical cord matrix or adipose tissue or amniotic fluid claim 1 , or urine.3. Process according to characterized in that it comprises mesenchymal stem cells frozen or fresh immobilized on inert supports.4. Process according to characterized in that it comprises hematopoietic stem/progenitor cells isolated from umbilical cord blood claim 1 , bone marrow claim 1 , mobilized peripheral blood and fetal liver.5. Process according to characterized in that it comprises mesenchymal stem cells of human origin.6. Process according to characterized in that it comprises mesenchymal stem cells and hematopoietic stem cells from the same donor.7. Process according to claim 1 , ...

CELL MICROSHEET, SYRINGE CONTAINING THE CELL MICROSHEET, AND PRODUCTION AND USE OF THE CELL MICROSHEET

Cell microsheets are formed from a culture of cells. The cell microsheets has a size that can pass through an injection needle with a certain thickness. The cell microsheets can be produced on a surface of a cell cultureware. A stimulus-responsive polymer is immobilized on the surface having small divisions of the cell cultureware. The cell microsheets are suitable for minimally invasive treatment. 1. A cell microsheet that is formed from a culture of cells and is capable of passing through an injection needle.2. The cell microsheet set forth in claim 1 , wherein the injection needle is an 18G or thinner injection needle.3. The cell microsheet set forth in claim 1 , having an area of 20 mmor less.4. The cell microsheet set forth in claim 1 , wherein the cell microsheet is usable for cartilage tissue repair.5. The cell microsheet set forth in claim 1 , wherein the cells are derived from cartilage tissue.6. The cell microsheet set forth in claim 5 , where the cartilage tissue is of an animal with polydactyly.7. The cell microsheet set forth in claim 1 , wherein the cells are derived from stem cells.8. The cell microsheet set forth in claim 7 , wherein the stem cells include pluripotent stem cells claim 7 , embryonic stem cells claim 7 , or somatic stem cells.9. The cell microsheet set forth in claim 7 , wherein the stem cells comprise iPS cells.10. A syringe containing the cell microsheet set forth in .11. A method of producing cell microsheets formed from a culture of cells claim 1 , comprising cultivating the cells on a surface of a cell cultureware to yield the cell microsheets claim 1 , a stimulus-responsive polymer being immobilized on the surface of the cell cultureware claim 1 , the surface having small divisions.12. The method set forth in claim 11 , wherein the small divisions each have an area of 20 mmor less.13. A method of administering the cell microsheet set forth in to an animal by injection. The present invention relates to a cell microsheet, a syringe ...

METHODS OF GROWING AND PREPARING STEM CELLS AND METHODS OF USING THE SAME

The present invention relates to methods of improving stem cell delivery to a subject in need thereof and kits designed to assist in such. The methods comprise interchangeably allowing or promoting cell growth in conditions that permit three-dimensional growth, such as with a bioreactor, utilizing allogeneic, autologous or xenogeneic cells, mixing the cells with platelet rich plasma that is autologous, allogeneic or xenogeneic to the subject, and site specific delivery of between about three and ten million activated stem cells per kilogram of the subject receiving the treatment. 1. A method of treating a site specific injury in a subject comprising administration of a suspension of stem cells to a site in need thereof , wherein the suspension comprises about 3-ten million activated stem cells per kilogram of the subject intravenously or about 5 to 10 million cells intra-articularly and further wherein the stem cells are derived from a 3D cell culture.2. The method of claim 1 , wherein the stem cells comprise autologous claim 1 , allogeneic or xenogeneic cells incubated in a bioreactor prior to administration to the subject.3. The method of claim 1 , wherein the suspension further comprises a bio-compatible matrix.4. The method of claim 1 , further comprising mixing the stem cells with autologous platelet rich plasma prior to administration to the subject.5. The method of claim 4 , wherein the autologous PRP is photo bio-stimulated.6. The method of claim 1 , further comprising mixing the suspension with autologous adipose-derived stem cells (ADSCs) isolated from the subject prior to administration.7. The method of claim 6 , wherein the autologous ADSCs are previously activated by PRP and/or photo-biostimulation.8. A method of treating a patient claim 6 , comprising administering a suspension of stem cells grown to the patient claim 6 , wherein the suspension of stem cells are derived from a 3D cell culture and contacted with platelet rich plasma and/or photo bio- ...

CELL CULTURE AUXILIARY AGENT AND CELL CULTURE MEDIUM USING THE SAME

A cell culture auxiliary agent and a cell culture medium using the same are provided. The cell culture auxiliary agent is formed by attaching each coordination peptide having cell affinity to two side ends of a polyoxyethylene polyoxypropylene ether block copolymer through anhydride monomers. 1. A cell culture auxiliary agent comprising a structural formula represented as follow: P-L-S-L-P; wherein in the structural formula , S is a substrate , each L is a linker , and each P is a peptide; and wherein the substrate is a polyoxyethylene polyoxypropylene ether block copolymer , the linkers are each independently an anhydride monomer , and an amino acid sequence of the peptides are each independently selected from a group consisting of glycine-arginine-glycine-aspartate , arginine-glycine-aspartate , arginine-glutamate-aspartate-valine , leucine-aspartate-valine , tyrosine-isoleucine-glycine-serine-arginine , proline-aspartate-serine-glycine-arginine , isoleucine-lysine-valine-alanine-valine , and arginine-asparagine-isoleucine-alanine-glutamate-isoleucine-isoleucine-lysine-aspartate-alanine.2. The cell culture auxiliary agent according to claim 1 , wherein the substrate is Pluronic® F-127 claim 1 , and the anhydride monomer is maleic anhydride claim 1 , succinic anhydride claim 1 , or 4-methacryloxyethyl trimellitic anhydride.3. The cell culture auxiliary agent according to claim 2 , wherein the anhydride monomer is maleic anhydride claim 2 , and the amino acid sequence of the peptide is glycine-arginine-glycine-aspartate.4. The cell culture auxiliary agent according to claim 2 , wherein the anhydride monomer is 4-methacryloxyethyl trimellitic anhydride claim 2 , and the amino acid sequence of the peptide is glycine-arginine-glycine-aspartate.5. A cell culture medium claim 2 , which comprises 0.5-5 wt % of a cell culture auxiliary agent including a structural formula represented as follow: P-L-S-L-P; wherein in the structural formula claim 2 , S is a substrate claim 2 ...

SYSTEMS AND METHODS FOR CULTURING CELLS IN SUSPENSION

A method of culturing adherent cells in suspension is provided that includes culturing adherent cells on a first substrate in a first suspension, harvesting the adherent cells from the first substrate, and transfecting the harvested adherent cells using electro-poration. The method also includes, after the step of transfecting, suspending the transfected adherent cells in a second suspension. A dissolution process for dissolving the second microcarrier particle to harvest the cells or cell products is also provided. This dissolution process includes adding a chelator, such as EDTA, to the second suspension for a predetermined time to separate the cells from the second microcarrier; and isolating the cells or cell products from a remainder of the second suspension after the predetermined time. The dissolution process is performed without enzymes such as pectinase or protease. 1. A method of culturing adherent cells in suspension , comprising:culturing adherent cells on a first substrate in a first suspension;harvesting adherent cells from the first substrate;transfecting the harvested adherent cells using electroporation;after the step of transfecting, suspending the transfected adherent cells in a second suspension.2. The method of claim 1 , wherein claim 1 , after electroporation claim 1 , the cells are recovered on a second substrate in suspension or in another suspension format.3. The method of claim 1 , the method further comprising harvesting the cells or products of the cells from the second suspension.4. The method of claim 1 , wherein the first substrate comprises a first microcarrier particle.5. The method of claim 1 , wherein the transfected adherent cells are suspended in the second suspension on a second substrate.6. The method of claim 5 , wherein the second substrate comprises a second microcarrier particle.7. The method of claim 6 , the method further comprising a dissolution process for dissolving the second microcarrier particle to harvest the cells ...

BLASTOID, CELL LINE BASED ARTIFICIAL BLASTOCYST

The invention relates to a method for making an at least double layered cell aggregate and/or an artificial blastocyst, and/or a further-developed blastoid termed blastoid, by forming a double layered cell aggregate from at least one trophoblast cell and at least one pluripotent and/or totipotent cell, and culturing said aggregate to obtain an artificial blastocyst. This artificial blastocyst has a trophectoderm-like tissue that surrounds a blastocoel and an inner cell mass-like tissue. The cell aggregate can be formed from toti- or pluripotent stem cell types, or induced pluripotent stem cell types, in combination with trophoblast stem cells. Formation of a blastoid can be achieved by culturing the cell aggregate in a medium preferably comprising one or more of a Rho/ROCK inhibitor, a Wnt pathway modulator, a PKA pathway modulator, a PKC pathway modulator, a MAPK pathway modulator, a STAT pathway modulator, an Akt pathway modulator, a Tgf pathway modulator and a Hippo pathway modulator. The invention further relates to a method for growing an at least double layered cell aggregate into an artificial blastocyst, and into a further-developed blastoid, a fetus or a live animal. The invention further pertains to an in vitro cell culture comprising the mentioned compounds and/or cell aggregates. 1. An in vitro method of making an at least double layered cell aggregate or a blastoid , the method comprising:forming an initial cell aggregate by combining at least one trophoblast cell and at least one pluripotent and/or totipotent cell; wherein the inner cell layer comprises inner cells which descend from the at least one pluripotent and/or totipotent cell and are capable of forming an embryo, and', 'wherein the outer cell layer comprises outer cells which descend from the at least one trophoblast cell and are capable of forming at least a trophectoderm., 'culturing the initial cell aggregate in a culture medium to obtain an at least double layered cell aggregate comprising ...

Culture Substrate, Method for Manufacturing Culture Substrate, and Culturing Method and Culturing Device for Stem Cell

The present disclosure provides a culture substrate for culturing stem cells, the culture substrate including a surface portion having: soft regions that extend side by side along a plurality of directions intersecting each other; and a plurality of stiff regions compartmented by the soft regions, wherein in the surface portion, the stiff regions have acute angle parts protruding toward the soft regions, and the cells can be deformed into a shape that is accommodated within the region of the stiff regions. 1. A culture substrate for culturing stem cells , soft regions that extend side by side along a plurality of directions intersecting each other; and', 'a plurality of stiff regions compartmented by the soft regions,, 'the culture substrate comprising a surface portion havingwherein in the surface portion, the stiff regions have acute angle parts protruding toward the soft regions, andthe stem cells can be deformed into a shape that is accommodated within the region of the stiff regions.2. The culture substrate according to claim 1 , wherein the stiff regions have a higher compressive modulus of elasticity than the soft regions.3. The culture substrate according to claim 1 , wherein the acute angle parts exhibit a chamfering shape claim 1 , and the radius of curvature thereof is 50 μm or less.4. The culture substrate according to claim 1 , wherein at least one of the plurality of stiff regions has a triangular shape.5. The culture substrate according to claim 1 , wherein the area of each of the stiff regions is 5 claim 1 ,000 to 13 claim 1 ,000 μm.6. The culture substrate according to claim 1 , wherein the compressive modulus of elasticity of the stiff regions is 10 or more times the compressive modulus of elasticity of the soft regions.7. The culture substrate according to claim 1 , wherein the compressive modulus of elasticity of the stiff regions is 30 kPa or higher.8. The culture substrate according to claim 1 , wherein the soft regions include a ...

Methods for Preparing Therapeutically Active Cells Using Microfluidics

The present invention is directed to the use of microfluidics in the preparation of cells and compositions for therapeutic uses. 1. A method for treating a patient suffering from a disease , comprising:a) obtaining a crude fluid composition comprising target cells and platelets;b) performing a size based separation on the crude fluid composition using a microfluidic device so as to reduce the ratio of platelets to target cells in the composition by at least 50%;c) genetically engineering target cells obtained from step b) to produce engineered cells;d) administering the engineered target cells to said patient;wherein the target cells are not centrifuged before or during the method.2. The method of claim 1 , wherein the crude fluid composition is blood claim 1 , an apheresis sample or a leukapheresis sample.3. The method of claim 2 , wherein the target cells are T cells and the engineered target cells are CAR T cells.4. The method of claim 3 , wherein the disease is cancer.5. The method of claim 4 , wherein claim 4 , in step b) claim 4 , the ratio of platelets to target cells is reduced by at least 80%.6. The method of claim 3 , wherein genetic engineering comprises transfecting or transducing the T cells.7. The method of claim 3 , wherein no more than four hours elapse from the time that the obtaining of the crude fluid composition comprising T cells is completed until T cells are transfected or transduced.8. The method of claim 3 , wherein the microfluidic device comprises:a) at least one channel extending from a sample inlet to one or more fluid outlets, wherein the channel is bounded by a first wall and a second wall opposite from the first wall;b) an array of obstacles disposed in the channel in a manner such that, when the crude fluid composition is applied to an inlet of the device and fluidically passed through the channel, T cells in the composition flow to one or more collection outlets where an enriched product is collected, and platelets flow to one more ...

CELL CULTURE DEVICE AND METHODS

A method of vascularising a cell aggregate on a microfluidic device, microfluidic cell culture devices comprising perfusable vascular networks and kits and assays using the microfluidic cell culture devices are described. The microfluidic devices comprise one or more capillary pressure barriers allowing for formation of an extracellular matrix gel within a confined area of the network, in which cells can be cultured for different uses. 188-. (canceled)89. A cell culture device comprising a microfluidic network , the microfluidic network comprising:a microfluidic layer comprising a base, a microfluidic channel and a cover;an organoid compartment extending into the microfluidic layer through an aperture in the cover and in fluid communication with the microfluidic channel; anda capillary pressure barrier substantially aligned with the aperture and dividing the microfluidic network into a first sub-volume comprising the organoid compartment and a second sub-volume comprising at least a part of the microfluidic channel.90. The cell culture device according to claim 89 , wherein the organoid compartment comprises a well having sidewalls that claim 89 , in an in use orientation claim 89 , extend vertically to the microfluidic layer.91. The cell culture device according to claim 89 , wherein the capillary pressure barrier is located on the base of the microfluidic layer substantially opposite the aperture.92. The cell culture device according to claim 89 , wherein the capillary pressure barrier defines at least in part a surface of the organoid compartment on the base of the microfluidic layer and is configured to confine a fluid to the first sub-volume.93. The cell culture device according to claim 89 , wherein the microfluidic network comprises a second capillary pressure barrier located on the base of the microfluidic layer substantially opposite and aligned with the aperture.94. The cell culture device according to wherein the second capillary pressure barrier further ...

Tumor model for breast cancer cell migration studies and related methods

A method for creating a tumor model includes encapsulating cancer cells in a first solution, disposing the first solution on a spacer, cross-linking the first solution and creating one or more high stiffness constructs, disposing a second solution around the one or more high stiffness constructs, and cross-linking the second solution and creating a low stiffness matrix surrounding the one or more low stiffness constructs.

Methods for Preparing Therapeutically Active Cells Using Microfluidics

The present invention is directed to the use of microfluidics in the preparation of cells and compositions for therapeutic uses. 1273-. (canceled)274. A method of producing Chimeric Antigen Receptor (CAR) T cells , comprising:a) obtaining a crude fluid composition comprising T cells; i) at least one channel extending from a sample inlet to one or more fluid outlets, wherein the channel is bounded by a first wall and a second wall opposite from the first wall;', 'ii) an array of obstacles arranged in rows in the channel, each subsequent row of obstacles being shifted laterally with respect to a previous row, and wherein said obstacles are disposed in a manner such that, when the crude fluid composition is applied to an inlet of the device and fluidically passed through the channel, T cells in the composition flow to one or more collection outlets where an enriched product is collected, and cells, or particles that are in the crude fluid composition and that are of a different size than the T cells, flow to one more waste outlets that are separate from the collection outlets;, 'b) performing Deterministic Lateral Displacement (DLD) on the crude fluid composition using a microfluidic device comprisingc) genetically engineering the T cells in the enriched product obtained in step b) to produce the chimeric antigen receptors (CARs) on their surface.275. The method of claim 274 , wherein said crude fluid composition is an apheresis product or leukapheresis product obtained from blood from a patient and wherein claim 274 , when the crude fluid composition is applied to an inlet of the device and fluidically passed through the channel claim 274 , T cells in the composition flow to one or more collection outlets where an enriched product is collected claim 274 , and red blood claim 274 , platelets or other particles that are in the crude fluid composition and that are of a different size claim 274 , flow to one more waste outlets that are separate from the collection outlets ...

DOUBLE TUBULAR STRUCTURES

The present invention relates to a method of culturing and/or monitoring epithelial cells using a microfluidic cell culture system comprising a microfluidic channel network. In the method epithelial cells are lined, in the microfluidic cell culture system by cells of mesenchymal origin. The cells may form a tubular or tube-like structure, i.e. a.tube in a tube. The method allows for improved epithelial models suitable for a wide variety of applications, including but not limited to high-throughput screening and analysis of epithelium in health and disease. 1. A method of culturing and/or monitoring epithelial cells using a microfluidic cell culture system comprising a microfluidic channel network , the method comprising a1) using an aqueous medium; or', 'a2) using a gel precursor and allowing the gelprecursor to gelate in the microfluidic channel network thereby occupying at least part of the microfluidic channel network;, 'a) introducing mesenchymal cells in the microfluidic channel network, wherein the mesenchymal cells are introduced in the microfluidic channel network'}b) in case of step a1), and preferably in case of step a2), allowing the mesenchymal cells to proliferate and/or differentiate, preferably until at least part of the microfluidic channel network is covered with mesenchymal cells;c) introducing epithelial cells in the microfluidic channel network comprising the mesenchymal cells; andd) allowing the epithelial cells to proliferate and/or differentiate, preferably until at least part of the microfluidic channel network is covered with epithelial cells and/or until at least part of the mesenchymal cells is covered with epithelial cells.2. The method of wherein a gel precursor is introduced in the microfluidic channel network and allowing the gelprecursor to gelate in the microfluidic channel network thereby occupying at least part of the microfluidic channel network.3. The method of wherein the gel is patterned claim 1 , preferably by use of a ...

ENGINEERED MICROGELS

Microparticles containing heparin or a heparin-like polymer and a biocompatible polymer are described. The heparin or the heparin-like polymer and the biocompatible polymer can be indirectly linked together by a coupling agent, which can have a structure represented by Formula I, (A)(R)(D), wherein A is a bond or a moiety that can form a bond with the heparin or the heparin-like polymer, D is a bond or a moiety that can form a bond with the biocompatible polymer, R is a linker for A and D, and p and q are from 1 to 25. Methods of making the microparticles include mixing a first solution of the heparin or the heparin-like polymer and a second solution of the biocompatible polymer, to form a mixture, and adding the mixture to an oil and a surfactant and homogenizing the mixture to form a water-in-oil emulsion. Compositions of these microparticles are also described. 1. A microparticle , comprising: heparin or a heparin-like polymer coupled to a biocompatible polymer , wherein the microparticle has an average diameter of from about 2 to about 30 μm.2. The microparticle of claim 1 , wherein the heparin or the heparin-like polymer is coupled to the biocompatible polymer by a coupling agent.3. The microparticle of claim 1 , wherein the coupling agent is represented by Formula I:{'br': None, 'sub': p', 'q, '(A)(R)(D)\u2003\u2003Formula I'}whereinA is a bond or a moiety which is bonded to the heparin or the heparin-like polymer,D is a bond or a moiety which is bonded to the biocompatible polymer,R is a linker for A and D,p and q are integers from 1 to 25.4. The microparticle of claim 3 , wherein A claim 3 , for each occurrence claim 3 , independently includes a moiety bonded to the heparin or the heparin-like polymer.5. The microparticle of claim 3 , wherein the moiety bonded to the heparin or the heparin-like polymer is formed from a Michael addition reaction claim 3 , nucleophilic substitution claim 3 , electrophilic substitution claim 3 , condensation reaction claim 3 , ...

Method for culturing mdck cells

The present invention relates to a cloned MDCK cell showing an expansion factor of 4.5 or more when cultured using a microcarrier and a method of culturing the MDCK cell, a method of growing a virus using the method of culturing the MDCK cell, and a cloned MDCK cell showing an expansion factor of 4.5 or more when cultured using a microcarrier.

Microcarriers for stem cell culture and fabrication thereof

A method for manufacturing polycaprolactone microcarriers is disclosed together with uses of the microcarriers.

MICROFLUIDIC THREE-DIMENSIONAL OSTEOCYTE NETWORK RECONSTRUCTED WITH MICROBEADS AS SCAFFOLD

A bed of microbeads is used as a foundation for reconstructing a three-dimensional osteocyte network by culturing osteocytes within the bed. The osteocytes are cultured such that they form a network among the microbeads that is capable of simulating the osteocyte network of natural bone. The osteocytes are cultured in a microfluidic device adapted for the purpose. 1. A method of culturing osteocytes in a microfluidic chamber , comprising the steps of:mixing a plurality of cells with a plurality of microbeads, thereby forming a mixture including cells and microbeads, the cells including one or both of a plurality of osteocytes and a plurality of pre-osteocytes;adding a portion of the mixture to a microfluidic chamber, thereby forming a bed of microbeads with cells distributed among the microbeads;allowing the cells of the bed to attach to the microbeads of the bed; andperfusing the bed with a culture medium, whereby osteocytes of the bed remain as osteocytes and pre-osteocytes of the bed develop into osteocytes.2. The method of claim 1 , wherein said perfusing step is performed such that one or both of the osteocytes of the bed and the pre-osteocytes of the bed form a network of osteocytes among the microbeads of the bed.3. The method of claim 1 , wherein the microbeads of the bed have diameters such that the bed has an interstitial space between adjacent ones of the microbeads of the bed claim 1 , the interstitial space having a size such that only one osteocyte or pre-osteocyte occupies the interstitial space.4. The method of claim 1 , wherein the cells are selected from a species of animal claim 1 , and the microbeads have diameters that are approximately the same size as a typical distance between osteocytes in a living animal of the species.5. The method of claim 1 , wherein the plurality of microbeads includes biphasic calcium phosphate.6. The method of claim 2 , wherein the culture medium includes a biologically-active substance claim 2 , and said method ...

In vitro method for culturing stem cells

There is provided a method for culturing a stem cell in vitro. The method comprises providing a substrate surface coated with a coating comprising a molecule having a catechol moiety or a polymer thereof; and growing a stem cell on said coated substrate surface in a growth medium.

MATERIALS AND METHODS FOR EXPANSION OF STEM CELLS

The subject invention concerns novel and translatable materials and methods for expansion of stem cells, such as mesenchymal stem cells (MSC), that significantly improve translational success of the cells in the treatment of various conditions, such as stroke. The subject invention utilizes cell self-aggregation as a non-genetic means to enhance their therapeutic potency in a microcarrier bioreactor. The subject invention integrates a cell aggregation process in a scalable bioreactor system. In one embodiment of the method, thermally responsive microcarriers (TRMs) are utilized in conjunction with a bioreactor system. Cells are cultured in a container or vessel in the presence of the TRMs wherein cells adhere to the surface of the TRMs. Once cells are adhered to the TRMs they can be cultured at a suitable temperature for cell growth and expansion, e.g., at about 37° C. After a period of time sufficient for cell growth and expansion on the TRMs, the cell culture temperature is reduced so that the cells detach from the TRMs. The detached cells are allowed to form cell clusters that are then cultured under conditions such that the clusters aggregate to form 3D aggregates. The 3D aggregates can be collected and treated to dissociate the cells (e.g., using enzymatic treatment, such as trypsinization). Dissociated cells can then be used for transplantation in methods of treatment or for in vitro characterization and study. 1. A method for expanding a stem cell , wherein said method comprises culturing stem cells in a bioreactor system in the presence of a thermally responsive microcarrier (TRM) , wherein stem cells adhere to the surface of said TRM; growing the adhered stem cells for a sufficient period of time for the stem cells to increase in numbers; detaching the stem cells from the TRM by reducing the culture temperature to a critical solution temperature that results in said adhered cells detaching from the surface of said TRM; providing said detached cells ...

Polymeric Carriers and Methods

Provided are methods of controlling disassociation of cells from a carrier, compositions, and methods of collecting cells. The methods of controlling disassociation of cells from a carrier may include contacting a polymeric carrier with one or more digesting agents to disassociate at least a portion of a plurality of cells from the polymeric carrier. The polymeric carrier may be crosslinked with a crosslinker including at least one of a redox sensitive moiety, a UV light sensitive moiety, a pH sensitive moiety, and a temperature sensitive moiety. 1. A method of controlling disassociation of cells from a carrier , the method comprising:providing a polymeric carrier and a plurality of cells adhered to the polymeric carrier; andcontacting the polymeric carrier with one or more digesting agents to disassociate at least a portion of the plurality of cells from the polymeric carrier;wherein the polymeric carrier is crosslinked with a crosslinker comprising at least one redox sensitive moiety.2. The method of claim 1 , wherein the at least one redox sensitive moiety comprises a disulfide bond.13. A method of controlling disassociation of cells from a carrier claim 1 , the method comprising:providing a polymeric carrier and a plurality of cells adhered to the polymeric carrier; andcontacting the polymeric carrier with one or more digesting agents to disassociate at least a portion of the plurality of cells from the polymeric carrier;wherein the polymeric carrier is crosslinked with a crosslinker comprising at least one of a UV light sensitive moiety, a pH sensitive moiety, and a temperature sensitive moiety.14. The method of claim 13 , wherein the crosslinker comprises the UV light sensitive moiety claim 13 , and the UV light sensitive moiety is a photoreversibly dimerizable moiety or a photocleavable moiety.19. The method of claim 14 , wherein the crosslinker comprises the photocleavable moiety claim 14 , and the photocleavable moiety comprises an o-nitrobezene based ...

MICROCARRIER PERFUSION CULTURING METHODS AND USES THEREOF

Provided herein are methods of culturing a mammalian cell and various methods that utilize these culturing methods. 1110.-. (canceled)111. A method for testing a manufacturing process for making a recombinant protein , the method comprising:providing a shake tube containing an adherent Chinese hamster ovary (CHO) cell containing a nucleic acid encoding a recombinant protein disposed in a first liquid culture medium, wherein the first liquid culture medium occupies about 10% to about 30% of the volume of the shake tube and contains a plurality of microcarriers at a concentration of about 1.0 g/L to about 15.0 g/L;incubating the shake tube for a period of time at about 32° C. to about 39° C. and with a rotary agitation of about 130 revolutions per minute (RPM) to about 150 RPM;{'sup': '6', 'after about the first 48 to 96 hours of the period of time, continuously or periodically removing a first volume of the first liquid culture medium and adding to the first liquid culture medium a second volume of a second liquid culture medium, wherein the first and second volumes are about equal, and the method achieves a viable cell density of greater than 2.0×10cells/mL in the first liquid culture medium or a combination of the first and second liquid culture medium at some point during the period of time;'}detecting the recombinant protein in the cell or in the first and/or second culture medium; andcomparing the amount of recombinant protein present in the adherent CHO cell or in the first and/or second culture medium to a reference level of recombinant protein.112. The method of claim 111 , wherein the reference level of recombinant protein is a level of recombinant protein produced using a different culturing method.113. The method of claim 112 , wherein the different culturing method utilizes a different first or second liquid culture medium claim 112 , a different adherent CHO cell claim 112 , a different temperature claim 112 , a different level of agitation claim 112 , a ...

RELEASE-MATRICES FOR CONTROLLED RELEASE OF MATERIALS INTO A SURROUNDING MEDIUM

The present invention refers to release matrices which include a material that can be released in a controlled way or with predefined kinetics into a surrounding medium, reaction vessels comprising said release matrices, and release system further comprising a medium which is able to dissolve the embedded material. The present invention further refers to the use of release matrices in several applications and the production method for such matrices. The present invention also relates to a method that al lows control of the release rate of materials from polymer matrices with several factors. 115-. (canceled)16. A release-matrix for controlled release of a nutrient , inducer or pH controlling agent into a medium , comprising a cross-linked polymer system based on a polysiloxane , a reactive polysiloxane and a nutrient , inducer or pH controlling agent embedded therein , wherein the particle size of the nutrient , inducer or pH controlling agent is from 1-20 μm.17. The release-matrix according to claim 16 , wherein the nutrient claim 16 , inducer or pH controlling agent is present in crystalline form or as a powder.18. The release-matrix according to claim 16 , wherein the nutrient claim 16 , inducer or pH controlling agent is distributed homogeneously in the release-matrix.19. The release-matrix according to claim 16 , wherein the release rate of the nutrient claim 16 , inducer or pH controlling agent into the medium is between 0.1-25 g/L*24 ĥ-1.20. A method for controlling the release rate of a nutrient claim 16 , inducer or pH controlling agent from a release-matrix according to into a surrounding medium claim 16 , wherein the release rate is controlled byi. the type of polysiloxane and/orii. an additive added to the polysiloxane, and/oriii. the degree of cross-linking of the polymer system, and/oriv. the size of particles/drops of nutrient, inducer or pH controlling agent, and/orv. the weight ratio of the polymer system to nutrient, inducer or pH controlling ...

3D Printing Of A Cellularised Scaffold

The present invention relates to a process for producing a droplet assembly, which droplet assembly comprises a plurality of droplets, wherein each of said droplets comprises: an aqueous medium comprising a hydrogel compound; and one or more biological cells disposed in the aqueous medium, which process comprises: generating, in a bulk hydrophobic medium, a plurality of droplets, wherein each of said droplets comprises: an aqueous medium comprising a hydrogel compound; and one or more biological cells disposed in the aqueous medium. The invention also relates to a droplet assembly comprising a plurality of droplets, wherein each of said droplets comprises: (i) an aqueous medium comprising a hydrogel compound; (ii) one or more biological cells disposed in the aqueous medium; and (iii) an outer layer of amphipathic molecules around the surface of the aqueous medium, wherein at least one droplet in the droplet assembly contacts at least one other droplet in the droplet assembly forming a layer of amphipathic molecules as an interface between contacting droplets.

The Treatment of Protein Aggregation Diseases

Compositions, methods and systems for the treatment of a protein aggregation disease including, but not limited to Alzheimer's disease (AD), Parkinson's disease (PD), Dementia with Lewy Bodies (DLB), Huntington's disease (HD), Amylotrophic lateral sclerosis (ALS, which results from degeneration of the upper and lower motor neurones and affects the voluntary muscle system), Progressive Supranuclear Palsy (PSP), Type 2 Diabetes and Multiple systems atrophy (MSA).

METHODS FOR CONTROLLED INDUCTION OF BIOENGINEERED NEUROEPITHELIAL TISSUES AND 3-D NEUROEPITHELIAL TUBES

Described herein are methods, compositions, and kits for directed differentiation of human pluripotent stem cells, neuromesodermal progenitors, and neural stem cells into bioengineered elliptical neuroepithelial tissues and bioengineered neuroepithelial tubes that contain a single rosette of polarized neuroepithelial cells and have microscale cellular organization similar to that of an in vivo developing human neural tube. 1. A method of producing a biomimetic elliptical neuroepithelial tissue having a singular rosette structure in vitro , the method comprising:(a) seeding human pluripotent stem cells (hPSCs) in the presence of a Rho kinase inhibitor onto a micropatterned substrate that is capable of biomimetic neural morphogenesis of cells cultured thereon, wherein the micropatterned substrate comprises at least two circular bounded regions connected by a cell-adhesive bridge;(b) culturing the seeded cells of step (a) on the micropatterned substrate for a first culture period of about one to two days in the presence of a pluripotency maintenance medium to obtain a first cell aggregate, wherein the pluripotency maintenance medium comprises a Rho kinase inhibitor; and(c) culturing the cells obtained in step (b) for a second culture period of about 3 to about 6 days under adherent culture conditions in a neural differentiation base medium,whereby a biomimetic elliptical neuroepithelial tissue having a singular rosette structure is obtained, wherein the tissue comprises polarized neuroepithelial cells and has a microscale cellular organization similar to that of a transverse section of an in vivo developing human neural tube.2. The method of claim 1 , wherein each of the at least two circular bounded regions has a diameter of about 100 μm to about 300 μm.3. The method of claim 1 , wherein the cell-adhesive bridge has a length of about 25 μm to about 125 μm claim 1 , and has a width of about 10 μm to about 50 μm.4. The method of claim 1 , wherein the hPSCs are seeded ...

Microgels and microtissues for use in tissue engineering

The present invention features microgels and microtissues for use in tissue engineering. Featured is a microencapsulation device for making microgels and/or microtissues via an emulsion technology. Also featured are methods of making higher ordered structures that mimic in vivo tissue structures. Methods of us are also featured.

Platelet-Targeted Microfluidic Isolation of Cells

Methods and systems for isolating platelet-associated nucleated target cells, e.g., such as circulating epithelial cells, circulating tumor cells (CTCs), circulating endothelial cells (CECs), circulating stem cells (CSCs), neutrophils, and macrophages, from sample fluids, e.g., biological fluids, such as blood, bone marrow, plural effusions, and ascites fluid, are described. The methods include obtaining a cell capture chamber including a plurality of binding moieties bound to one or more walls of the chamber, wherein the binding moieties specifically bind to platelets; flowing the sample fluid through the cell capture chamber under conditions that allow the binding moieties to bind to any platelet-associated nucleated target cells in the sample to form complexes; and separating and collecting platelet-associated nucleated target cells from the complexes. 122-. (canceled)23. A two-stage microfluidic system for isolating platelet-associated nucleated target cells from a sample fluid comprising: a microchannel having an inlet, a waste outlet, a product outlet, and', 'an array of microposts arranged between the inlet and the outlets, wherein the microposts are arranged in rows and spaced apart by a distance that enables red blood cells and unbound platelets to flow through the device to a waste outlet and to cause platelet-associated nucleated target cells to be laterally displaced by the array of microposts to a product outlet, wherein the microposts in each subsequent row are offset laterally from microposts in a previous row by a distance less than the spacing between the microposts within the row;, 'a first chamber comprising'} a microchannel having an inlet and an outlet, wherein fluid flows from the inlet to the outlet through the microchannel, and binding moieties fixed to at least one internal surface of the microchannel, wherein the binding moieties specifically bind to platelets; and', 'a fluid connection between the product outlet of the first chamber and ...

Biomarker Detection Methods and Systems and Kits for Practicing Same

Aspects of the present disclosure include methods that include co-culturing a cell and a microparticle that includes a capture ligand, in a culture medium under conditions in which a biomarker produced by the cell is bound by the capture ligand. Such methods may further include detecting (e.g., by flow or mass cytometry) complexes that include the microparticle, the capture ligand, the biomarker, and a detection reagent. The methods may further include determining the proportion or number of cells among a heterogeneous cell population that produced the biomarker and/or the level of biomarker secreted by such cells. Compositions, systems and kits are also provided. 1. A method , comprising: a cell; and', 'a microparticle comprising a capture ligand,, 'co-culturingin a culture medium under conditions in which a biomarker produced by the cell is bound by the capture ligand.2. The method according to claim 1 , wherein the biomarker bound by the capture ligand is secreted from the cell.3. The method according to claim 2 , wherein the biomarker is selected from the group consisting of: a cytokine claim 2 , an immunoglobulin claim 2 , a hormone claim 2 , a growth factor claim 2 , an enzyme claim 2 , a protease claim 2 , a protein claim 2 , an allergen claim 2 , a peptide claim 2 , a nucleic acid claim 2 , a drug claim 2 , a cluster differentiation (CD) molecule claim 2 , a tumor marker claim 2 , a receptor claim 2 , and combinations thereof.4. The method according to claim 3 , wherein the biomarker is a cytokine selected from the group consisting of: an interferon claim 3 , a chemokine claim 3 , an interleukin claim 3 , a lymphokine claim 3 , a tumor necrosis factor claim 3 , and combinations thereof.5. The method according to claim 1 , wherein the method comprises stimulating the cell to secrete the biomarker.6. The method according to claim 5 , wherein stimulating the cell to secrete the biomarker comprises adding a stimulant to the culture medium.7. The method according ...

MICROCARRIERS FOR CELL CULTURE

A cell culture microcarrier bead is proposed. The microcarrier bead comprises a bead body having its surface flecked with plasmonic nanoparticles. In a second aspect, the invention relates to a cell culture reactor, containing a cell culture medium and the proposed microcarrier beads. A third aspect of the invention concerns a method for observing living cells on such microcarrier beads. Yet a further aspect of the invention relates to a method for packing nanoparticles on a carrier body. 1. A cell culture microcarrier bead , comprising a bead body having a surface , the surface flecked with plasmonic nanoparticles , the bead body having a diameter in the range from 50 μm to 1 mm. the bead body comprising a swollen or unswollen hydrogel.21. The microcarrier bead as claimed in , wherein the plasmonic nanoparticles comprise plasmonic metal nanoparticles.3. The microcarrier bead as claimed in claim 1 , wherein the plasmonic metal nanoparticles comprise gold nanoparticles.4. The microcarrier bead as claimed in claim 1 , wherein the plasmonic nanoparticles are stabilized with a capping agent claim 1 , e.g. citrate.5. The microcarrier bead as claimed in claim 1 , wherein the plasmonic nanoparticles are spherical.6. The microcarrier bead as claimed in claim 1 , wherein the plasmonic nanoparticles have a diameter in the range from 2 to 200 nm.7. The microcarrier bead as claimed in claim 1 , wherein the ratio of the diameter of the bead body to the diameter of the plasmonic nanoparticles lies in the range from 250 to 25000.8. A cell culture reactor claim 1 , containing a cell culture medium and microcarrier beads as claimed in .9. A method for observing living cells on a microcarrier bead claim 1 , the microcarrier bead comprising a bead body having a surface claim 1 , the surface being flecked with plasmonic nanoparticles claim 1 , the bead body having a diameter in the range from 50 μm to 1 mm. the bead body comprising a swollen or unswollen hydrogel claim 1 , the method ...

Methods and compositions for t cell activation

Among the various aspects of the present disclosure is the provision of methods, synthetic DC, and compositions for T cell activation. The present disclosure provides for synthetic dendritic cells (DCs), methods of generating synthetic dendritic cells (DCs), methods of generating T cell-encapsulated gelatin microspheres and microcapsules, methods of activating T cells using synthetic DCs, methods for expanding T cells against individualized antigen-specific mutational antigens using synthetic DCs, and methods of treating a chronic disease (e.g., HIV, HPV) or cancer using the synthetic DCs.

GENETICALLY ENGINEERED HUMAN FETAL LIVER NICHE AS PLATFORM FOR BIOMANUFACTURING OF HEMATOPOIETIC STEM CELLS

The present disclosure relates to methods for expanding populations of hematopoietic stem cells (HSCs) using a genetically engineered human fetal liver niche and compositions of purified ex vivo expanded HSCs. Also provided herein are methods of using such expanded HSC cell populations for clinical applications including allogeneic hematopoietic stem cell transplantation and for drug discovery and modeling human liver development. 1. A method for ex vivo expansion of human hematopoietic stem cells (HSCs) , the method comprising{'sup': +', '+', '+, '(a) contacting a cell population comprising human hematopoietic stem cells (HSCs) to a synthetic fetal liver organoid comprising mesenchymal-like cells, CD34-expressing (CD34) endothelial-like cells, desmin-expressing (DES) stellate-like cells, and CEBPα hepatocyte-like cells; and'}(b) culturing the contacted organoid under conditions that promote HSC proliferation for about 3 to about 10 days, whereby an expanded population comprising CD34+ human HSCs is obtained.2. The method of claim 1 , wherein the expanded population comprises at least 3-fold more CD34+ HSCs than the cell population of step (a).3. The method of claim 1 , further comprising obtaining the cell population of step (a) from a human subject.4. The method of claim 1 , wherein the cell population comprising HSCs is derived from umbilical cord blood or bone marrow.5. The method of claim 1 , wherein the synthetic fetal liver organoid constitutes a coating on a solid carrier.6. The method of claim 5 , wherein the carrier is a particulate support.7. The method of claim 1 , further comprising administering the expanded population of CD34+ human HSCs to a subject from whom the original cell population comprising HSCs is obtained.8. The method of claim 1 , wherein the contacted organoid is cultured in a medium selected from IMDM medium and APEL medium.9. The method of claim 8 , wherein the culture medium is supplemented with one or more of Stem Cell Factor (SCF) ...

CELL CULTURE ARTICLE AND METHODS THEREOF

A cell culture article, including: a substrate comprising a polygalacturonic acid compound selected from at least one of: pectic acid; partially esterified pectic acid having a degree of esterification from 1 to 40 mol %, or salts thereof; and an adhesion polymer on the surface of the polygalacturonic acid compound. A method of making and using the article are also disclosed. 1. A cell culture article , comprising:a substrate comprising a polygalacturonic acid compound selected from at least one of:pectic acid;partially esterified pectic acid having a degree of esterification from 1 to 40 mol %, or salts thereof; andan adhesion polymer on the surface of the polygalacturonic acid compound.2. The article of wherein the polygalacturonic acid compound is covalently cross linked claim 1 , ionically cross linked claim 1 , or mixtures thereof.3. The article of claim 1 , wherein the partially esterified pectic acid comprises an alkyl carboxy ester having an alkyl group having from 1 to 10 carbon atoms.4. The article of claim 1 , wherein the adhesion polymer on the surface of the polygalacturonic acid compound comprises a polypeptide.5. The article of claim 1 , wherein the adhesion polymer on the surface of the polygalacturonic acid compound comprises a polymer having a conjugated polypeptide.7. The article of claim 1 , wherein the adhesion polymer is present in an amount of from 0.1 to 30 weight % based on the total weight of the article claim 1 , or based on the total weight of the polygalacturonic acid compound or compounds selected.8. The article of claim 1 , wherein the adhesion polymer promotes the attachment of anchorage dependent live cells to the substrate.9. The article of claim 1 , wherein the substrate comprises a microcarrier particle.10. A method for harvesting cultured cells claim 1 , comprising:{'claim-ref': {'@idref': 'CLM-00001', 'claim 1'}, 'culturing cells on the surface of the article of ; and'}contacting the cultured cells with a mixture of pectinase ...

MICROBEADS FOR CELL CULTURE AND METHOD OF MONITORING CELL CULTURE USING THE SAME

Disclosed are microbeads for cell culture and a method of monitoring cell culture using the same. More particularly, each of the microbeads for cell culture according to an embodiment of the present invention include a core and a surface modification layer formed on a surface of the core. By using the method of monitoring cell culture with the microbeads for cell culture according to an embodiment of the present invention, cell culture may be carried out in highly scaled-up dimension and easily monitored. 1. A microbead for cell culture , comprising:a core; anda surface modification layer formed on a surface of the core.2. The microbead according to claim 1 , further comprising a metal coating layer formed between the core and the surface modification layer.3. The microbead according to claim 1 , wherein the microbead has a specific gravity of 0.90 to 1.00.4. The microbead according to claim 1 , wherein the microbead has a spherical or disc shape.5. The microbead according to claim 4 , wherein the microbead has a diameter of 10 μm to 800 μm.6. The microbead according to claim 1 , wherein the core comprises at least any one selected from the group consisting of glass claim 1 , silica claim 1 , plastic such as polystyrene (PS) claim 1 , polyethylene (PE) claim 1 , polypropylene (PP) claim 1 , and biocompatible polymers such as poly lactic acid (PLA) claim 1 , poly L-lactic acid (PLLA) claim 1 , poly(glycolic acid) (PGA) claim 1 , poly(lactic-co-glycolic acid) (PLGA) claim 1 , and poly-caprolactone (PCL).7. The microbead according to claim 1 , wherein the surface modification layer comprises at least any one selected from the group consisting of gelatin claim 1 , collagen claim 1 , hyaluronic acid claim 1 , chondroitin sulfate claim 1 , alginate claim 1 , chitosan claim 1 , aminopropylsiloxane claim 1 , poly-dopamine claim 1 , poly-L-lysine claim 1 , RGD peptide claim 1 , and graphene.8. The microbead according to claim 2 , wherein the metal coating layer comprises at ...

T-cell expansion method and uses

The invention provides a method for the expansion of anti-tumor T-cells, comprising the steps of: a) providing a phagocytosable particle, having one or more tumor neoantigenic constructs tightly associated thereto, wherein the tumor neoantigenic construct comprises an amino-acid sequence comprising at least one mutated amino acid known or suspected to be associated with a cancer in a subject, or a mutated or non-mutated amino-acid sequence known or suspected to be expressed in a cancer cell in the subject; b) providing a viable antigen-presenting cell; c) contacting the particle with the antigen-presenting cell in vitro under conditions allowing phagocytosis of the particle by the antigen-presenting cell; d) providing a T-cell sample comprising viable T-cells from the subject; e) contacting the T-cell sample with the antigen-presenting cell contacted with the particle in vitrounder conditions allowing specific activation of anti-tumor T-cells in response to antigen presented by the antigen-presenting cell. The invention also provides tumor neoantigenic constructs as defined in the specification.

MICROCARRIER PERFUSION CULTURING METHODS AND USES THEREOF

Provided herein are methods of perfusion culturing an adherent mammalian cell using a shake flask and a plurality of microcarriers, and various methods that utilize these culturing methods. 1. A method of optimizing a manufacturing process of producing a recombinant protein , the method comprising:providing a shake flask containing a mammalian cell disposed in a first liquid culture medium, wherein the first liquid culture medium occupies about 20% to about 30% of the volume of the shake flask and contains a plurality of microcarriers at a concentration of about 1.0 g/L to about 15.0 g/L;incubating the shake flask for a period of time at about 32° C. to about 39° C. and with a rotary agitation of about 85 revolutions per minute (RPM) to about 125 RPM; and after about the first 48 to 96 hours of the period of time, continuously or periodically removing a first volume of the first liquid culture medium and adding to the first liquid culture medium a second volume of a second liquid culture medium, wherein the first and second volumes are about equal;detecting the recombinant protein in the cell or in the first and/or second culture medium;comparing the amount of recombinant protein present in the cell or in the first and/or second culture medium to a reference level of recombinant protein produced by a different method; andidentifying and removing or altering in a manufacturing process any culture components or parameters that are associated with a decrease in the amount of recombinant protein produced as compared to the reference level, or identifying and adding to a manufacturing process any culture components or parameters that are associated with an increase in the amount of recombinant protein produced as compared to the reference level.2. The method of claim 1 , wherein the first volume of the first liquid culture medium is substantially free of the microcarriers.3. The method of claim 1 , wherein the first liquid culture medium occupies about 25% to about 30% ...

CURVATURE-DEFINED CONCAVE AND CONVEX PDMS SURFACES FOR USE IN CELL AND TISSUE CULTURING AND IN OTHER SURFACE AND INTERFACE APPLICATIONS

The present disclosure provides a method of fabricating curvature-defined (C-D) or shape-defined (S-D) concave and convex polydimethylsiloxane (PDMS) surfaces and a method of fabricating C-D or S-D convex and concave gel surfaces for use in cell and tissue culturing and in other surface and interface applications, and provides a method of using C-D or S-D convex and concave surfaces with varying curvatures to direct cell attachment, spreading, and migration. 1. A method of fabricating curvature-defined (C-D) or shape-defined (S-D) concave and convex surfaces for use in cell and tissue culturing and in other surface and interface applications , comprising:(1) embedding rigid C-D or S-D convex microstructures on a solidified first material layer of a sufficient rigidity through the polymerization or solidification process to form this solidified first material layer of a sufficient rigidity, and then the exposed C-D or S-D concave surfaces being obtained by carefully-removing these embedded rigid C-D or S-D convex microstructures from this solidified first material layer of a sufficient rigidity, wherein the curvatures of the obtained exposed C-D or S-D concave surfaces are same to those of the C-D or S-D convex surfaces of the corresponding removed rigid convex microstructures that generated these exposed C-D or S-D concave surfaces, and wherein, the sufficient rigidity of a solidified material layer means that (same below), this solidified material layer is rigid enough or the elastic moduli of this solidified material layer is large enough so that, the permanent deformations on the to-be-exposed concave surface of this solidified material layer induced by the possible significant pulling and pushing forces between an embedded rigid C-D or S-D convex microstructure and this solidified material layer during the removing process of this embedded rigid convex microstructure, and the shape variations of the exposed concave surface of this solidified material layer ( ...

Thermoresponsive Microcarrier System and Uses Thereof

There is provided a polymeric microsphere comprising a thermally responsive monomer crosslinked with a functional group monomer, wherein the functional group monomer comprises at least one of a carboxylic acid functional group or an amine functional group. The thermally responsive monomer is preferably N-isopropylacrylamide (NIPAM), and the microspheres preferably comprise a coating of polymerized catecholamines (e.g. DOPA). There is also provided a method of preparing the polymeric microsphere and uses of the polymeric microsphere in culturing, harvesting, or expanding stem cells or stromal cells. Preferably, the cells, e.g. hMSCs (human mesenchymal stem/stromal cells), are expanded or harvested in serum-free and xeno-free medium. 1. A polymeric microsphere comprising a thermally responsive monomer crosslinked with a functional group monomer , wherein the functional group monomer comprises at least one of a carboxylic acid functional group or an amine functional group.2. The polymeric microsphere of claim 1 , wherein the thermally responsive monomer is selected from the group consisting of N-isopropylacrylamide claim 1 , N claim 1 ,N-diethylacrylamide claim 1 , 2-(dimethylamino)ethyl methacrylate claim 1 , N claim 1 ,N-dimethylacrylamide claim 1 , acrylamide claim 1 , 2-(diethylamino)ethyl acrylate claim 1 , 2-(acryloyloxyethyl) trimethylammonium chloride claim 1 , poly(vinylcaprolactame) claim 1 , polyvinyl methyl ether claim 1 , poly(hydroxyethylmethacrylate) claim 1 , 4-hydroxybutyl acrylate claim 1 , 2-hydroxyethyl methacrylate claim 1 , 3-hydroxypropyl methacrylate claim 1 , 2-carboxyethyl acrylate claim 1 , 2-carboxyethyl acrylate oligomers claim 1 , and poly(ethylene glycol) methacrylate.3. The polymeric microsphere of claim 1 , wherein the carboxylic acid functional group monomer is selected from the group consisting of methacrylic acid claim 1 , acrylic acid claim 1 , and 2-carboxyethyl acrylate.4. The polymeric microsphere of claim 1 , wherein the amine ...

CULTURE MEDIUM COMPOSITION AND METHOD OF CULTURING CELL OR TISSUE USING THEREOF

The present invention provides a culture method of cells and/or tissues including culturing cells and/or tissues in a suspended state by using a medium composition wherein indeterminate structures are formed in a liquid medium, the structures are uniformly dispersed in the solution and substantially retain the cells and/or tissues without substantially increasing the viscosity of the solution, thus affording an effect of preventing sedimentation thereof, and the like 1. A medium composition comprising (i) deacylated gellan gum or a salt thereof , and (ii) a polysaccharide other than deacylated gellan gum or a salt thereof , wherein the polysaccharide other than deacylated gellan gum or a salt thereof is at least one kind selected from the group consisting of xanthan gum , alginic acid , carageenan , diutan gum , and a salt thereof.2. The medium composition according to claim 1 , wherein the polysaccharide other than deacylated gellan gum or a salt thereof is alginic acid or a salt thereof.3. The medium composition according to claim 1 , wherein the deacylated gellan gum or salt thereof and the polysaccharide other than deacylated gellan gum or a salt thereof are present at a concentration which allows cells or a tissue to be cultured in suspension standing culture.4. The medium composition according to claim 1 , wherein the composition has a viscosity of not more than 8 mPa·s at 37° C.5. The medium composition according to claim 1 , wherein the polysaccharide other than deacylated gellan gum or a salt thereof is at least one kind selected from the group consisting of methylcellulose claim 1 , locust bean gum claim 1 , and a salt thereof.6. The medium composition according to claim 5 , wherein the polysaccharide other than deacylated gellan gum or a salt thereof is methylcellulose or a salt thereof.7. The medium composition according to claim 5 , wherein the deacylated gellan gum or salt thereof and the polysaccharide other than deacylated gellan gum or a salt ...

DRYING FORMULATON FOR HYDROGEL MICROCARRIERS

A method of making a cell culture article is provided. The method includes forming a microcarrier from a microcarrier composition comprising a polygalacturonic acid compound or an alginic acid compound, infiltrating the microcarrier with a drying formulation to form an infiltrated microcarrier, and drying the infiltrated microcarrier to form a dried microcarrier, wherein the drying formulation comprises at least one of a saccharide and a monovalent cation. 1. A method of making a cell culture article , the method comprising:forming a microcarrier from a microcarrier composition comprising a polygalacturonic acid compound or an alginic acid compound;infiltrating the microcarrier with a drying formulation to form an infiltrated microcarrier; anddrying the infiltrated microcarrier to form a dried microcarrier, wherein the drying formulation comprises at least one of a saccharide and a monovalent cation.2. The method according to claim 1 , further comprising sterilizing the dried microcarrier to form a sterilized dried microcarrier.3. The method according to claim 2 , wherein sterilizing the dried microcarrier comprises exposing the dried microcarrier to gamma radiation.4. The method according to claim 1 , further comprising rehydrating the microcarrier.5. The method according to claim 1 , wherein infiltrating the microcarrier with a drying formulation comprises soaking the microcarrier in a solution of the drying formulation.6. The method claim 1 , wherein infiltrating the microcarrier with a drying formulation comprises simultaneously spraying the microcarrier composition and the drying formulation.7. The method according to claim 1 , wherein the drying formulation comprises 1 to 50 wt. % saccharide.8. The method according to claim 1 , wherein the drying formulation comprises 10 to 500 mM monovalent cation.9. The method according to claim 1 , wherein the drying formulation comprises 1 to 50 wt. % saccharide and 10 to 500 mM monovalent cation.10. The method according ...

Methods for Preparing Therapeutically Active Cells Using Microfluidics

The present invention is directed to the use of microfluidics in the preparation of cells and compositions for therapeutic uses. 1. A method for preparing cells for treating a patient with cancer , comprising the steps: i) performing a size based separation using a microfluidic device to produce an enriched product in which, compared to the sample, the percentage cells that are platelets has been reduced; and', 'ii) in addition to the size based separation, performing an affinity based separation;, 'a) purifying T cells from a sample comprising leukocytes and platelets, wherein the leukocytes comprise T cells and wherein the T cells are purified byb) after the purification of step a), activating and expanding the T cells to produce a composition in which the percentage of T cells that are central memory T cells has increased compared to the percentage of T cells that are central memory T cells in the sample;c) genetically engineering activated T cells to comprise therapeutic benefit in the treatment of said patient's cancer.2. The method of claim 1 , wherein claim 1 , in step c) claim 1 , the activated T cells are genetically engineered to comprise modified cell surface receptors of therapeutic benefit in the treatment of said patient's cancer.3. The method of claim 2 , wherein claim 2 , the modified cell surface receptors of therapeutic benefit are chimeric antigen receptors (CARs).4. The method of claim 1 , wherein the sample is obtained by apheresis5. The method of claim 1 , wherein the sample is obtained by leukapheresis.6. The method of claim 1 , wherein the platelets in the enriched product of paragraph a)ii) are depleted by at least 80% compared to the sample and/or there are no more than 5 platelets per leukocyte in the enriched product.7. The method of claim 1 , wherein claim 1 , the genetically engineered T cells are collected by transferring them into a pharmaceutical composition for administration to a patient.8. The method of claim 7 , wherein cells are ...

DRYING FORMULATION FOR HYDROGEL MICROCARRIERS

A method of making a cell culture article is provided. The method includes forming a microcarrier from a microcarrier composition comprising a polygalacturonic acid compound or an alginic acid compound, infiltrating the microcarrier with a drying formulation to form an infiltrated microcarrier, and drying the infiltrated microcarrier to form a dried microcarrier, wherein the drying formulation comprises at least one of a saccharide and a monovalent cation. 113-. (canceled)14. A cell culture article comprising:a polygalacturonic acid compound or an alginic acid compound; anda drying formulation comprising at least one of a saccharide and a monovalent cation,wherein the cell culture article is free of water.15. The cell culture article according to claim 14 , wherein the drying formulation comprises 1 to 50 wt. % saccharide.16. The cell culture article according to claim 14 , wherein the drying formulation comprises 0.5 to 20 wt. % monovalent cation.17. The cell culture article according to claim 14 , wherein the drying formulation comprises 1 to 50 wt. % saccharide and 0.5 to 20 wt. % monovalent cation.18. The cell culture article according to claim 14 , wherein the drying formulation comprises a non-volatile liquid material.19. The cell culture article according to claim 18 , wherein the non-volatile liquid material is selected from the group consisting of DMSO and a low molecular weight polyethylene glycol.20. The cell culture article according to claim 14 , wherein the polygalacturonic acid compound or the alginic acid compound is cross-linked with calcium.21. The cell culture article of claim 14 , wherein the cell culture article comprises a microcarrier comprising the polygalacturonic acid compound or the alginic acid compound claim 14 , wherein the drying formulation is infiltrated into the microcarrier.22. The cell culture article of claim 14 , wherein the drying formulation comprises 10 to 500 mM monovalent cation.23. The cell culture article of claim 14 , ...

DIGESTIBLE SUBSTRATES FOR CELL CULTURE

A cell culture article is provided. The cell culture substrate includes a polygalacturonic acid compound selected from at least one of: pectic acid or salts thereof, and partially esterified pectic acid having a degree of esterification from 1 to 40 mol % or salts thereof. The polygalacturonic acid compound is crosslinked with a divalent cation and the divalent cation concentration ranges from 0.5 to 2 g/1 of the substrate. 1. A cell culture article , comprising:a substrate comprising a polygalacturonic acid compound selected from at least one of:pectic acid or salts thereof, andpartially esterified pectic acid having a degree of esterification from 1 to 40 mol % or salts thereof,wherein the polygalacturonic acid compound is crosslinked with a divalent cation and the divalent cation concentration ranges from 0.5 to 2 g/l of the substrate.2. The article of claim 1 , wherein the substrate is spherical or substantially spherical.3. The article of claim 2 , wherein the substrate comprises a diameter of 10 to 500 micrometers.4. The article of claim 1 , wherein a plurality of the cell culture articles comprise a coefficient of variation of less than 20%.5. The article of claim 1 , wherein a plurality of the cell culture articles comprise a coefficient of variation of less than 10%.6. The article of claim 2 , wherein a plurality of the cell culture articles comprise size spread Δd5-d95 of less than 25 micrometers claim 2 , wherein d5 is a diameter that is larger than the diameters of 5% of the plurality of the cell culture articles claim 2 , wherein d95 is a diameter that is larger than the diameters of 95% of the plurality of the cell culture articles claim 2 , and wherein Δd5-d95 is the difference between d95 and d5.7. The article of claim 2 , wherein a plurality of the cell culture articles comprise size spread Δd10-d90 of less than 20 micrometers claim 2 , wherein d10 is a diameter that is larger than the diameters of 10% of the plurality of the cell culture articles ...

CULTURE METHOD AND CELL CLUSTER

The present invention provides a culture method for culturing, in recesses (), a population including two or more cells including a cell derived from a stem cell and a mesenchymal cell. The cell derived from a stem cell is a cell obtained by differentiating a stem cell in vitro. The cell is a cell of one or more types selected from the group consisting of an endodermal cell, an ectodermal cell, and a mesodermal cell. The population is cultured in the recesses () together with a vascular cell or a secretor factor. Each recess () includes a space in which cells are movable. When a volume of the space is represented by V mmand the number of mesenchymal cells seeded in the space is represented by N, V is 400 or less and N/V is in a range from 35 to 3000. 1. A culture method comprising:three-dimensionally culturing a population including two or more cells in a predetermined area, the population including a cell derived from a stem cell and a mesenchymal cell, the cell derived from a stem cell being a cell of one or more types selected from the group consisting of an undifferentiated endodermal cell, an undifferentiated ectodermal cell, and an undifferentiated mesodermal cell, whereinthe area is formed of a space in which cells are movable, and{'sup': '3', 'when the space has a volume of V mmand the number of the mesenchymal cells seeded in the space is represented by N, the V is equal to or less than 400 and N/V is in a range from 35 to 3000.'}2. The culture method according to claim 1 , further comprising performing the three-dimensional culture of the population together with at least one of: a vascular cell; a factor autonomously secreted from a vascular cell; and or a factor secreted from a vascular cell due to the presence of both a vascular cell and a mesenchymal cell.3. The culture method according to claim 1 , wherein a ratio of the number of the mesenchymal cells to a total number of cells used for culture is 0.5% or more but less than 5%.4. The culture method ...

ENGINEERED TISSUE CONSTRUCTS

A modular engineered tissue construct includes a plurality of fused self-assembled, scaffold-free, high-density cell aggregates. At least one cell aggregate includes a plurality of cells and a plurality of biocompatible and biodegradable nanoparticles and/or microparticles that are incorporated within the cell aggregates. The nanoparticles and/or microparticles acting as a bulking agent within the cell aggregate to increase the cell aggregate size and/or thickness and improve the mechanical properties of the cell aggregate as well as to deliver bioactive agents. 1. A method of forming an engineered tissue construct , the method comprising:providing undifferentiated and/or substantially differentiated progenitor cells;combining the undifferentiated and/or substantially differentiated progenitor cells with a plurality of the nanoparticles and/or microparticles so that the nanoparticles and/or microparticles are dispersed and suspended with the undifferentiated and/or substantially differentiated progenitor cells in a culture medium; andculturing the suspension of nanoparticles and/or microparticles and undifferentiated and/or substantially differentiated progenitor cells in a well having a defined shape to form a self-assembled cell aggregate, the shape of the self-assembled cell aggregate being defined by surfaces of the well.2. The method of claim 1 , wherein the population of undifferentiated and/or substantially differentiated progenitor cells are autologous claim 1 , allogeneic claim 1 , or a combination thereof.3. The method of claim 1 , wherein the population of undifferentiated and/or substantially differentiated progenitor cells comprises at least one of chondrogenic cells claim 1 , mesenchymal stem cells claim 1 , endothelial cells claim 1 , and/or smooth muscle cells.4. The method of claim 1 , wherein the step of providing the undifferentiated and/or substantially differentiated progenitor cells includes:isolating undifferentiated and/or substantially ...

METHOD AND CONTAINER FOR CULTURING UNDIFFERENTIATED INDUCED PLURIPOTENT STEM CELL

Provided are a method of culturing an induced pluripotent stem cell and a method of forming a spheroid of the induced pluripotent stem cell. According to an embodiment of the present inventive concept, the induced pluripotent stem cell may have improved stemness. 1. A container for culturing an induced pluripotent stem cell , the container having a nanostructure array.2. The container of claim 1 , wherein the nanostructure array has a plurality of nanocolumns that are spaced at random intervals on a substrate.3. The container of claim 2 , wherein the length of each nanocolumn is in a range of about 0.5 μm to about 1.5 μm.4. The container of claim 2 , wherein the density of nanocolumns in the nanostructure array is in range of about 150/100 μmto about 1 claim 2 ,000/100 μm(the number of nanocolumns/area of nanostructure array).5. The container of claim 1 , wherein the container is designed to culture the induced pluripotent stem cell in an undifferentiated condition under a feeder layer-free.6. The container of claim 1 , wherein the container is for forming a spheroid of the induced pluripotent stem cell.7. The container of claim 1 , wherein the nanostructure array is reusable.8. A method of culturing an induced pluripotent stem cell under an undifferentiated condition claim 1 , the method comprising:culturing an induced pluripotent stem cell in a nanostructure array.9. The method of claim 8 , wherein the nanostructure array has a plurality of nanocolumns that are spaced at random intervals on a substrate.10. The method of claim 8 , wherein the length of each nanocolumn in the nanostructure array is in a range of about 0.5 μm to about 1.5 μm.11. The method of claim 8 , wherein the density of nanocolumns in the nanostructure array is in range of about 150/100 μmto about 1 claim 8 ,000/100 μm(the number of nanocolumns/area of nanostructure array).12. A method of forming a spheroid of an induced pluripotent stem cell claim 8 , the method comprising:culturing an induced ...

METHODS AND COMPOSITIONS FOR SPINAL CORD CELLS

Described here are systems and methods for deriving both spinal motor neurons and brain microvascular endothelial cells from induced pluripotent stem cells using distinct methods and combining them in a chip format. Neurons cultured alone in chip microvolume displayed increased calcium transient function and chip-specific gene expression. When seeded with endothelial cells, interaction further enhanced neural function, elicited vascular-neural interaction, niche gene expression with enhanced in vivo-like signatures arising from the chip co-cultures. Development of novel media formulations further allow for improved readout of differentiation process, by eliminating additives that otherwise confound differentiation processes and resulting phenotypes. 1. A method of generating spinal neural progenitor cells (spNPCs) comprising:providing induced pluripotent stem cells (iPSCs);differentiating iPSCs into neural ectodermal cells by culturing in neural induction media; anddissociating neural ectodermal cells, replating on a cell culture substrate and further culturing the replated neural ectodermal cells in differentiation media to generate spNPCs.2. The method of claim 1 , wherein the iPSCs are cultured in neural induction media for a period of about 5-7 days.3. The method of claim 1 , wherein the replated neural ectodermal cells are cultured in differentiation media for a period of about 5-7 days.4. The method of claim 1 , wherein the neural induction media comprises one or more of: LDN193189 claim 1 , SB431542 claim 1 , and CHIR99021.5. The method of claim 1 , wherein the cell culture substrate is matrigel.6. The method of claim 1 , wherein the differentiation media comprises one or more of: ascorbic acid claim 1 , retinoic acid claim 1 , SAG.7. The method of claim 1 , further comprising freezing the spNPCs.8. A cryopreserved solution of spNPCs made by the method of .10. A cell culture media comprising:{'claim-ref': {'@idref': 'CLM-00009', 'claim 9'}, 'the cell culture ...

SYSTEMS AND METHODS FOR SCALABLE MANUFACTURING OF THERAPEUTIC CELLS IN BIOREACTORS

Systems and methods for scalable manufacturing of therapeutic cells in bioreactors are disclosed. Fluid dynamic considerations for scale in accordance with an implementation include a method of production of therapeutic cells grown on microcarriers or as cell aggregates in a suspension-based bioreactor includes depositing a suspension comprising cells suspended in a volume of culture fluid into a bioreactor and setting an agitation rate of a mixer disposed in the bioreactor. The method includes actuating the mixer at the set agitation rate to mix the suspension in the bioreactor. The suspension includes a plurality of turbulent eddies generated by the mixer. A magnitude of an energy dissipation rate (EDR) of at least approximately 60% of the turbulent eddies can be less than approximately 0.0015 m2/s3. 1. A method of scaling production of therapeutic cells grown on microcarriers or as cell aggregates in a suspension-based bioreactor , the method comprising:determining a target average energy dissipation rate (EDR) of turbulent eddies within a suspension including cells disposed in a small scale bioreactor;determining a small scale agitation rate to achieve the target average EDR in the small scale bioreactor;determining a large scale agitation rate to achieve the target average EDR in a large scale bioreactor, the large scale agitation rate being directly dependent on the small scale agitation rate;depositing a suspension comprising a plurality of cells suspended in a volume of culture fluid into the large scale bioreactor;setting an agitation rate of a mixer disposed in the large scale bioreactor to the large scale agitation rate; andactuating the mixer in the large scale bioreactor at the large scale agitation rate to mix the suspension with an average EDR approximately equal to the target average EDR.2. The method of claim 1 , wherein the average EDR comprises an average of a plurality of actual EDR data points within the volume of the suspension in the large ...

SYSTEMS AND METHODS FOR THE SEPARATION OF CELLS FROM MICROCARRIERS USING A SPINNING MEMBRANE

Methods and systems for processing suspensions of biological cells and microcarriers are disclosed. The biological cells are separated from the microcarriers by introducing the suspension into a spinning membrane separator whereby the biological cells pass through the membrane and the microcarriers do not pass through the membrane. 1. A method for separating biological cells from microcarriers in a suspension comprising:a) introducing a suspension of biological cells and microcarriers into a separation device comprising a relatively rotatable cylindrical housing and an internal member, wherein said cylindrical housing has an interior surface and said internal member has an exterior surface, said surfaces defining a gap therebetween, wherein one of said surfaces includes a porous membrane comprising pores sized to retain said microcarriers while allowing said biological cells to pass through said membrane; andb) withdrawing said biological cells from said separation device.2. The method of wherein said pore size is less than or equal to approximately 50 μm.3. The method of wherein said pore size is greater than or equal to approximately 20 μm.4. The method of comprising introducing said suspension of biological cells into said gap.5. The method of comprising introducing said suspension of biological cells from a container that is in fluid communication with said gap.6. The method of wherein said biological cells are introduced through an inlet in flow communication with said gap and said microcarriers are removed through an outlet in flow communication with said gap.7. The method of wherein said biological cells are adhered to the surface of said microcarriers during said introducing step.8. The method of further comprising introducing a cleaving agent into said container prior to introducing said biological cells into said separator.9. The method of wherein said microcarriers comprise polymeric claim 1 , coated beads.10. The method of wherein said membrane is made ...

METHOD FOR CULTURING PLURIPOTENT STEM CELLS

Provided is a method for efficiently culturing pluripotent stem cells with higher safety. The present invention relates to a method for culturing pluripotent stem cells, the method comprising culturing an isolated pluripotent stem cells in a pseudo-microgravity environment to proliferate the pluripotent stem cells while maintaining the pluripotent stem cells in an undifferentiated state, thereby forming and growing spheroids of the pluripotent stem cells; and a method for inducing differentiation of pluripotent stem cells by using the method. 1. A method for culturing induced pluripotent stem cells (iPS cells) , the method comprising: culturing isolated iPS cells in a pseudo-microgravity environment to proliferate the iPS cells while maintaining the iPS cells in an undifferentiated state , thereby forming and growing spheroids of the iPS cells; and performing one cycle of the steps of disrupting the resulting spheroids by passing the spheroid through a filter having a filter mesh size of 40 to 100 μm and culturing the disrupted spheroids in a pseudo-microgravity environment , thereby forming and growing spheroids , or repeating the cycle two or more times.2. (canceled)3. The method according to claim 1 , wherein the culturing is performed in the absence of a cell scaffold material.4. The method according to claim 1 , wherein the iPS cells are seeded at a cell density of 4×10to 6×10cells/cm.5. The method according to claim 1 , wherein the culturing is performed in the presence of an apoptosis inhibitor.6. The method according to claim 5 , wherein the apoptosis inhibitor is a ROCK inhibitor.7. The method according to claim 1 , wherein the pseudo-microgravity environment is an environment in which an object is subjected to a gravity corresponding to 1/10 to 1/100 of the earth's gravity in time average.8. The method according to claim 1 , wherein the pseudo-microgravity environment is obtainable by using a uniaxial rotary bioreactor capable of achieving a pseudo- ...

BIOPRESERVED STEM CELLS ON MICROCARRIERS

Disclosed herein are compositions and methods involving stem cells biopreserved on microcarriers, which can be thawed and expanded, all while maintaining their key attributes, such as their proliferative capacity, identity, functionality, and potency. Disclosed is a method for generating microcarriers-seeded-stem cells, as well as a post biopreservation procedure for thawing and inoculating bioreactors to achieve a rapid and scalable stem cells expansion. 1. A composition comprising biopreserved stem cells adhered to a microcarrier.2. The composition of claim 1 , wherein the biopreserved stem cells have a cell viability of at least 70% and retain the ability to expand at least 10 fold after being stored for at least 6 months at −200 to −20° C. after recovery.3. The composition of claim 2 , wherein the composition comprises a cryopreservative agent.4. The composition of claim 3 , wherein the cryopreservative agent comprises Dimethyl sulfoxide (DMSO).5. The composition of claim 1 , wherein the biopreserved stem cells have a cell viability of at least 70% and retain the ability to expand at least 10 fold after being stored for 1 to 14 days at 0 to 10° C. after recovery.6. The composition of claim 5 , wherein the composition comprises a biopreservative agent.7. The composition of claim 6 , wherein the biopreservative agent comprises HypoThermosol®.8. The composition of claim 2 , wherein the biopreserved stem cells maintain cell surface marker expression claim 2 , lineage differentiation potential claim 2 , and cell functionality after recovery.9. The composition of claim 1 , wherein the composition comprises about 1-50 cells per microcarrier.10. The composition of claim 1 , wherein the composition comprises the biopreserved stem cells at a concentration of 1×10to 5×10cells/ml.11. The composition of claim 1 , wherein cell-adhered microcarriers are stored in a vial claim 1 , bag claim 1 , cryovial claim 1 , or cryobag.12. The composition of claim 1 , wherein the stem ...

Culture medium composition and method of culturing cell or tissue using thereof

The present invention provides a culture method of cells and/or tissues including culturing cells and/or tissues in a suspended state by using a medium composition wherein indeterminate structures are formed in a liquid medium, the structures are uniformly dispersed in the solution and substantially retain the cells and/or tissues without substantially increasing the viscosity of the solution, thus affording an effect of preventing sedimentation thereof, and the like

Artificial cartilage and method for its production

Disclosed is a three-dimensional tissue culture, comprising chondrocytes in a biocompatible artificial matrix, having at least the following layers: a first layer located at or close to a surface of the matrix, wherein chondrocytes have a non-spherical shape and are arranged essentially in parallel to the surface along their longest dimension; and a second layer at least partially covered by the first layer wherein the mean sphericity of the chondrocytes of the second layer is higher than the mean sphericity of the chondrocytes of the first layer; and preferably a third layer at least partially covered by the second layer, wherein chondrocytes are arranged into columns extending into the matrix, wherein each column has at least two chondrocytes. Such a tissue culture may for instance be used as artificial cartilage in surgery. Also disclosed is a method to produce such a three-dimensional culture.

Microcarriers for Stem Cell Culture

We disclose a particle comprising a matrix coated thereon and having a positive charge, the particle being of a size to allow aggregation of primate or human stem cells attached thereto. The particle may comprise a substantially elongate, cylindrical or rod shaped particle having a longest dimension of between 50 μm and 400 μm, such as about 200 μm. It may have a cross sectional dimension of between 20 μm and 30 μm. The particle may comprise a substantially compact or spherical shaped particle having a size of between about 20 μm and about 120 μm, for example about 65 μm. We also disclose a method of propagating primate or human stem cells, the method comprising: providing first and second primate or human stem cells attached to first and second respective particles, allowing the first primate or human stem cell to contact the second primate or human stem cell to form an aggregate of cells and culturing the aggregate to propagate the primate or human stem cells for at least one passage. A method of propagating human embryonic stem cells (hESCs) in long term suspension culture using microcarriers coated in Matrigel or hyaluronic acid is also disclosed. We also disclose a method for differentiating stem cells.

Platelet-Targeted Microfluidic Isolation of Cells

Methods and systems for isolating platelet-associated nucleated target cells, e.g., such as circulating epithelial cells, circulating tumor cells (CTCs), circulating endothelial cells (CECs), circulating stem cells (CSCs), neutrophils, and macrophages, from sample fluids, e.g., biological fluids, such as blood, bone marrow, plural effusions, and ascites fluid, are described. The methods include obtaining a cell capture chamber including a plurality of binding moieties bound to one or more walls of the chamber, wherein the binding moieties specifically bind to platelets; flowing the sample fluid through the cell capture chamber under conditions that allow the binding moieties to bind to any platelet-associated nucleated target cells in the sample to form complexes; and separating and collecting platelet-associated nucleated target cells from the complexes.

Algae Beads

Algae-containing beads comprising a microorganism such as unicellular microalgae, an insoluble carbon source such as char, water, and a crosslinked organic matrix. The beads can further contain clay, such as kaolin. 1. An algal bead comprising unicellular microalgae , char , and a crosslinked organic matrix.2. The bead of claim 1 , further comprising a clay.3. The bead of claim 1 , wherein the organic matrix before crosslinking comprises a plurality of hydroxyl groups that react with a divalent cation to crosslink the matrix.4. The bead of claim 3 , wherein the organic matrix comprises a water soluble polysaccharide selected from the group consisting of alginate salts claim 3 , galactomannan claim 3 , gellan gum claim 3 , carrageenan claim 3 , agarose claim 3 , and mixtures thereof.5. The bead of claim 2 , wherein the clay comprises kaolin.6. The bead of claim 1 , wherein the char comprises a biochar.7. The bead of claim 6 , wherein the biochar is produced from algal biomass or rice hulls.8. The bead of claim 1 , further comprising soluble salts of cations selected from Na+ claim 1 , K+ claim 1 , Mg+2 claim 1 , Ca+2 claim 1 , Fe+3 claim 1 , Mn+2 claim 1 , Zn+2 claim 1 , Cu+2 claim 1 , and Co+2.9. The bead of claim 8 , comprising soluble salts of one or more anions selected from nitrates claim 8 , phosphates claim 8 , hydrogen phosphates claim 8 , dihydrogen phosphates claim 8 , sulfates claim 8 , chlorides claim 8 , EDTA claim 8 , carbonates claim 8 , bicarbonates claim 8 , and molybdates.10Chlorella, Spirulina, Dunaliella, Haematococcus, Crypthecodinium, Schizochytrium, Scenedesmus, Aphanizomenon, Arthrospira, Odontella, Isochrysis, Nannochloropsis, Tetraselmis, Phaeodactylum, Porphyridium,. The bead of claim 1 , wherein the microalgae are selected from the group consisting of and combinations thereof.11. The bead of claim 3 , wherein the divalent ion is selected from Be+2 claim 3 , Mg+2 claim 3 , Ca+2 claim 3 , Sr+2 claim 3 , and Ba+2.12. A method of making a bead ...

NOVEL MICROCARRIER BEADS

The invention relates to a novel microcarrier bead; a method for producing same; a therapeutic comprising said microcarrier bead and attached thereto or grown thereon at least one selected cell or tissue type; a method for making said therapeutic; and a method of treatment involving the use of said microcarrier bead or said therapeutic. 1. A microcarrier bead made from apatite and characterised by one or more , including any combination , of the following features:a) micrometre-sized;b) a regular porous structure;c) rough surface;d) substantially spherical;e) osteo-conductivity;f) chemical similarity to the mineral phase of natural bone; andg) high thermal stability permitting them to be easily sterilized.2. A microcarrier bead according to wherein said apatite is selected form the group comprising: hydroxyapatite claim 1 , silicon-substituted apatite claim 1 , silver-substituted apatite claim 1 , magnesium-substituted apatite and a stoichiometric apatite which is a synthetic apatite with a Ca/P atomic ratio that approaches 1.67.3. A microcarrier bead according to wherein said apatite is phase-pure.4. A microcarrier bead according to wherein said beads are between 100-800 μm diameter claim 1 , or 200-600 μm diameter claim 1 , or 400-500 μm diameter.5. A microcarrier bead according to wherein said beads have a regular pore size as observed by Scanning Electron Microscopy.6. A microcarrier bead according to wherein said beads can withstand temperatures up to 1500° C. for up to 10 hours.7. A microcarrier bead according to wherein said beads have osteogenic potency.8. A plurality of microcarrier beads according to any one of .9. A method for making microcarrier beads comprising:a) mixing apatite and alginate in a solution and allowing them to disperse to form a suspension;b) extruding said suspension drop-wise through a droplet device;{'sub': '2', 'c) exposing said extruded droplets to calcium chloride (CaCl) solution;'}{'sub': '2', 'd) washing said beads to remove said ...

Custom multiwell plate design for rapid assembly of photo-patterned hydrogels

The present invention provides a system for conservation and efficient use of energy through controlling and monitoring of devices. At least one processing controller connected to a sensor and a device, the processing controller configured to receive the ambient data from the sensor and operating parameters from the device; a user module configured to IO receive input parameters from a plurality of users; a central processing module, connected to the structure, the user module, and the admin module through wired and/or wireless connection, the central processing module configured to process the data received from the processing controller adapted in the zone of the structure and generate the optimum parameters for operating the device adapted in the zone to the structure.

Method of Coating Surfaces with Nanoparticles for Biological Analysis of Cells

A method of coating a surface with nanoparticles for biological analysis of cells that includes the steps of cleaning the surface with an oxidizing acid, treating the surface with an organosilane, coating the surface with nanoparticles, and then growing cells on the surface coated with the nanoparticles. The surface may be a glass surface, a silica-based surface, a plastic-based surface or a polymer-based surface. The nanoparticles may be gold-based nanomaterials. 1. A method of coating a surface with a nanoparticle for biological analysis of a cell , comprising the steps of:a. cleaning said surface with an oxidizing acid;b. treating said surface with an organosilane;c. coating said surface with said nanoparticle; andd. growing said cell on said surface coated with said nanoparticle.2. The method of claim 1 , wherein said surface is a glass surface claim 1 , a silica-based surface claim 1 , a plastic-based surface or a polymer-based surface.3. The method of claim 1 , wherein said nanoparticle comprises gold.4. The method of claim 1 , wherein said nanoparticle comprises a nanocage.5. The method of claim 1 , further comprising the step of conducting a biological analysis of said cell.6. The method of claim 5 , wherein said biological analysis comprises exposing said cell to radiation. This application claims the benefit of U.S. Provisional Application No. 62/630,575, entitled “Nanoparticle Coated Surface for Precision Nanobiology” and filed on Feb. 14, 2018. The complete disclosure of said patent application is hereby incorporated by reference.This invention was made with government support from grant no. 01A-Award 1457888 awarded by the National Science Foundation. The government has certain rights in the invention.Local thermal therapy is an attractive, emerging treatment modality for various biomedical conditions requiring sterilization of pathological tissue features (clots, plaques, tumors, or infections). Gold or iron oxide nanoparticles are under intense ...

Ferromagnetic cell and tissue culture microcarriers

A porous, collagen coated, ferromagnetic cell culture microcarrier, which is suitable for in vitro cell and tissue culture and which facilitates 3D multicellular construct generation. Also provided is a method for creating batches of microcarriers which have inserted within them magnetite (Fe3O4) in the presence of collagen, thus creating a microcarrier which becomes magnetic in nature when placed in a the presence of a magnetic field and which facilitates cellular adherence (via the collagen coating) for 3D construct development.

Topographical Templating Of Polymeric Materials Using Cellular Morphology

Substrates for influencing the organization, spreading or adhesion of a selected cell to induce or stimulate growth, differentiation on regeneration of the cell or of tissue constituting the cells are provided as well as methods of making such substrates and methods of using such substrates. 2. The method of claim 1 , further comprising the step of adding a guidance cue to the pre-polymer solution.3. The method of claim 1 , further comprising the step of coating the elastomeric polymer cell-templated substrate with a guidance cue after separating the elastomeric polymer cell-templated substrate from the cell template.4. The method of claim 1 , further comprising the step of forming the elastomeric polymer cell-templated substrate into a tube or channel having an interior surface.5. The method of claim 4 , wherein the features of the elastomeric polymer cell-templated substrate project exteriorly from the interior surface of the tube or channel.6. The method of claim 4 , wherein the features of the elastomeric polymer cell-templated substrate project interiorly from the interior surface of the tube or channel.7. The method of claim 1 , wherein the selected human cell is a Schwann cell claim 1 , an astrocyte claim 1 , or an oligodendrocyte.8. The method of claim 1 , wherein the elastomeric claim 1 , polymer cell-templated substrate is composed of a natural elastomeric polymeric gel claim 1 , a natural elastomeric polymeric solid claim 1 , a synthetic elastomeric polymeric gel claim 1 , or a synthetic elastomeric polymeric solid.9. The method of claim 1 , wherein the elastomeric polymer cell-templated substrate includes an alkylsiloxane claim 1 , a polylactic acid claim 1 , a poly(D claim 1 ,L-lactide) claim 1 , a copolymer of lactic acid and glycolic acid claim 1 , a copolymer of lactic acid and ε-aminocaproic acid claim 1 , a polyhydroxyalkanoate claim 1 , a polyester claim 1 , a polyglycolic acid claim 1 , a polycaprolactone claim 1 , a polydesoxazon claim 1 , a ...

PLURIPOTENT STEM CELL CULTURE ON MICRO-CARRIERS

The present invention is directed to methods for the growth, expansion and differentiation of pluripotent stem cells on micro-carriers. 1. A method for the propagation of human pluripotent stem cells comprising:a) Attaching a population of human pluripotent stem cells to a first volume of micro-carriers;b) Culturing the human pluripotent stem cells on the first volume of micro-carriers in a defined medium lacking a Rho kinase inhibitor,c) Removing the human pluripotent stem cells from the first volume of micro-carriers; andd) Attaching the population of human pluripotent stem cells to a second volume of micro-carriers.2. The method of claim 1 , wherein the steps of culturing claim 1 , removing and attaching the human pluripotent stem cells on micro-carriers are is repeated using subsequent volumes of micro-carriers.3. The method of claim 1 , wherein the first volume of micro-carriers is selected from the group consisting of dextran micro-carriers and polystyrene micro-carriers.4. The method of claim 1 , wherein the second volume of micro-carriers is selected from the group consisting of dextran micro-carriers and polystyrene micro-carriers.5. The method of claim 1 , wherein the human pluripotent stem cells are attached to the first volume of micro-carriers claim 1 , the second volume of micro-carriers claim 1 , or both in medium containing a Rho kinase inhibitor.6. The method of claim 5 , wherein the Rho kinase inhibitor is Y27632 or Glycyl-H 1152 dihydrochloride.7. The method of claim 6 , wherein the method comprises from about 1 μM to about 10 μM of the Rho kinase inhibitor Y27632.8. The method of claim 6 , wherein the method comprises from about 0.25 μM to about 5 μM of the Rho kinase inhibitor Glycyl-H 1152 dihydrochloride.9. The method of claim 1 , wherein the human pluripotent stem cells are removed from the first volume of micro-carriers claim 1 , the second volume of micro-carriers claim 1 , or both by enzymatic treatment.10. The method of claim 1 , wherein ...

METHODS OF PERFUSION CULTURING USING A SHAKE FLASK AND MICROCARRIERS

Provided herein are methods of perfusion culturing an adherent mammalian cell using a shake flask and a plurality of microcarriers, and various methods that utilize these culturing methods. 1. A method of testing the efficacy of a first or second liquid culture medium , a raw ingredient or supplement present in a first or second liquid culture medium , or a source of an adherent mammalian cell for use in a method of producing a recombinant protein , the method comprising:providing a shake flask containing an adherent mammalian cell disposed in a first liquid culture medium, wherein the first liquid culture medium occupies about 20% to about 30% of the volume of the shake flask and contains a plurality of microcarriers at a concentration of about 1.0 g/L to about 15.0 g/L;incubating the shake flask for a period of time at about 32° C. to about 39° C. and with a rotary agitation of about 85 revolutions per minute (RPM) to about 125 RPM; and{'sup': '6', 'after about the first 48 to 96 hours of the period of time, continuously or periodically removing a first volume of the first liquid culture medium and adding to the first liquid culture medium a second volume of a second liquid culture medium, wherein the first and second volumes are about equal, the method achieves a viable cell density of greater than 2×10cells/mL in the first liquid culture medium or a combination of the first liquid culture medium and the second liquid culture medium at some point during the period of time, and the adherent mammalian cell contains a nucleic acid encoding the recombinant protein;'}detecting the recombinant protein in the adherent mammalian cell or in the first and/or second culture medium;comparing the amount of recombinant protein present in the cell or in the first and/or second culture medium to a reference level of recombinant protein produced by a different method that uses one or more of a different first or second liquid culture medium, a different raw ingredient or ...

METHODS OF PREPARING HEMATOPOIETIC PROGENITOR CELLS IN VITRO

Disclosed are methods of preparing CD34+CD43+ hematopoietic progenitor cells (HPC) in vitro according to embodiments of the invention. Also disclosed are methods of differentiating CD34+CD43+ hematopoietic progenitor cells to hematopoietic lineage cells according to embodiments of the invention. Also disclosed are methods of treating or preventing a condition in a mammal, e.g., cancer, according to embodiments of the invention. 1. A method of preparing CD34+CD43+ hematopoietic progenitor cells (HPC) in vitro , the method comprising:(i) culturing source cells in a vessel;(ii) collecting the source cells from the vessel and preparing a single cell suspension of source cells;(iii) culturing the single cell suspension in a first three-dimensional (3D) culture and forming embryoid bodies (EBs) from the source cells in the first 3D culture;(iv) collecting the EBs, mixing the EBs with mesoderm lineage cells (MLC), and preparing a single cell suspension comprising EBs and MLC;(v) co-culturing the EBs and MLC in the single cell suspension in a second 3D culture and forming hematopoietic spheroids in the second 3D culture;(vi) collecting the hematopoietic spheroids from the second 3D culture and culturing the hematopoietic spheroids in a third 3D culture; and(vii) harvesting CD34+CD43+ hematopoietic progenitor cells from the hematopoietic spheroids.2. The method of claim 1 , wherein one or more of the first claim 1 , second claim 1 , and third 3D cultures is a hanging drop 3D culture claim 1 , a 3D microwell culture claim 1 , a 3D culture on a hydrophobic surface claim 1 , a rotational culture claim 1 , or a static 3D suspension culture.3. The method of claim 1 , wherein one or more of the first claim 1 , second claim 1 , and third 3D cultures is a bioreactor.4. The method of claim 1 , wherein (vi) comprises culturing the hematopoietic spheroids in xeno-free medium.5. The method of claim 4 , wherein the xeno-free medium comprises human platelet lysate.6. The method of claim 1 ...

MATERIALS AND METHODS FOR EXPANSION OF STEM CELLS

The subject invention concerns novel and translatable materials and methods for expansion of stem cells, such as mesenchymal stem cells (MSC), that significantly improve translational success of the cells in the treatment of various conditions, such as stroke. The subject invention utilizes cell self-aggregation as a non-genetic means to enhance their therapeutic potency in a microcarrier bioreactor. The subject invention integrates a cell aggregation process in a scalable bioreactor system. In one embodiment of the method, thermally responsive microcarriers (TRMs) are utilized in conjunction with a bioreactor system. Cells are cultured in a container or vessel in the presence of the TRMs wherein cells adhere to the surface of the TRMs. Once cells are adhered to the TRMs they can be cultured at a suitable temperature for cell growth and expansion, e.g., at about 37° C. After a period of time sufficient for cell growth and expansion on the TRMs, the cell culture temperature is reduced so that the cells detach from the TRMs. The detached cells are allowed to form cell clusters that are then cultured under conditions such that the clusters aggregate to form 3D aggregates. The 3D aggregates can be collected and treated to dissociate the cells (e.g., using enzymatic treatment, such as trypsinization). Dissociated cells can then be used for transplantation in methods of treatment or for in vitro characterization and study. 1. A method for expanding a stem cell , wherein said method comprises culturing stem cells in a bioreactor system in the presence of a thermally responsive microcarrier (TRM) , wherein stem cells adhere to the surface of said TRM and wherein said TRM is a microcarrier bead coated with or comprising a thermally responsive material selected from one or more of poly(N-isopropylacrylamide) (PNIPAAm) , poly(allylamine hydrochloride)-co-poly(N-isopropylacrylamide) , or poly(styrene sulfonate)-co-poly(N-isopropylacrylamide); growing the adhered stem cells for ...

Method for reducing drug-induced nephrotoxicity

A method for reducing renal tissue toxicity in a subject caused by a kidney damaging agent is disclosed. The method comprises administering to the subject: (i) a kidney damaging agent; and (ii) an inhibitor of glucose reabsorption.

An Implantable Construct, Methods of Manufacturing, and Uses Thereof

The present invention refers to a method of manufacturing an implantable construct comprising chondrogenically differentiated cells and one or more polycaprolactone (PCL) microcarriers, an implantable construct produced using said method, and uses of the implantable construct. The present invention also refers to a method of manufacturing an implantable construct comprising mesenchymal stromal cells and one or more polycaprolactone (PCL) microcarriers, an implantable construct produced using said method, and uses of the implantable construct. The present invention further refers to a method of treating a disease or disorder associated with cartilage and/or bone defect, the method comprises administering one or more cell-free polycaprolactone (PCL) microcarriers in a patient suffering from the disease or disorder. 1. A method of manufacturing an implantable construct comprising chondrogenically differentiated cells and one or more polycaprolactone (PCL) microcarriers , the method comprising:a) culturing mesenchymal stromal cells with one or more PCL microcarriers in a suspension culture in a mesenchymal stromal cells growth medium to allow the mesenchymal stromal cells to attach to the PCL microcarriers to form one or more mesenchymal stromal cells-PCL microcarrier complexes, wherein the suspension culture is agitated;b) harvesting the one or more mesenchymal stromal cells-PCL microcarrier complexes from the suspension culture in a) while the suspension culture is agitated;c) culturing the one or more mesenchymal stromal cells-PCL microcarrier complexes from b) under agitation-free and centrifugation-free conditions in the mesenchymal stromal cells growth medium;d) culturing the one or more mesenchymal stromal cells-PCL microcarrier complexes from c) under agitation-free and centrifugation-free conditions in a chondrogenic differentiation medium to enact differentiation of the mesenchymal stromal cells into chondrogenically differentiated cells.2. The method of claim ...

Filtered cell culture caps and cell culture methods

A bioreactor is provided herein. The bioreactor includes a vessel having a wall at least partially defining an interior compartment for receiving fluid, at least one port, and at least one cap configured to removably engage with the at least one port, the at least one cap comprising a filter material. A cell culture method is also provided herein which includes adding cells and cell growth medium to a vessel of a bioreactor and adding microcarriers to the vessel to form substantially confluent cells on the microcarriers. The cell culture method further includes washing the confluent cells, harvesting the confluent cells to form a solution containing the cells, and removing the solution containing the cells from the vessel by flowing the solution through a filter material in a cap removably engaged with at least one port of the bioreactor.

Manufacturing Process for Polysaccharide Beads

The invention discloses a method of manufacturing polysaccharide beads, comprising the steps of: i) providing a water phase comprising an aqueous solution of a polysaccharide; ii) providing an oil phase comprising at least one water-immiscible organic solvent and at least one oil-soluble emulsifier; iii) emulsifying the water phase in the oil phase to form a water-in-oil (w/o) emulsion; and iv) inducing solidification of the water phase in the w/o emulsion, wherein the organic solvent is an aliphatic or alicyclic ketone or ether. 1. A method of manufacturing polysaccharide beads , comprising the steps of:i) providing a water phase comprising an aqueous solution of a polysaccharide;ii) providing an oil phase comprising at least one water-immiscible organic solvent and at least one oil-soluble emulsifier;iii) emulsifying said water phase in said oil phase to form a water-in-oil (w/o) emulsion; andiv) inducing solidification of said water phase in said w/o emulsion,wherein said at least one organic solvent is an aliphatic or alicyclic ketone or ether.2. A method of manufacturing polysaccharide beads , comprising the steps of:i) providing a water phase comprising an aqueous solution of a polysaccharide;ii) providing an oil phase comprising at least one water-immiscible organic solvent and at least one oil-soluble emulsifier;iii) emulsifying said water phase in said oil phase to form a water-in-oil (w/o) emulsion; andiv) inducing solidification of said water phase in said w/o emulsion,{'sup': 1/2', '1/2', '1/2', '1/2', '1/2', '1/2, 'wherein said at least one organic solvent does not contain halogens and has Hansen solubility parameter values in the ranges of δD=15.0-18.5 MPa, δP=3.5-8.5 MPaand δH=4.0-5.5 MPa, or wherein said oil phase comprises a mixture of halogen-free water-immiscible organic solvents, said mixture having Hansen solubility parameter values in the ranges of δD=15.0-18.5 MPa, δP=3.5-8.5 MPaand δH=4.0-5.5 MPa.'}3. The method of wherein said at least one ...

TRYPSIN-FREE CELL STAMP SYSTEM AND USE THEREOF

The present invention relates to a trypsin-free cell stamp system and a use thereof. According to the present invention, an increase in the passage number of stem cells can be prevented compared with conventional methods of isolating cells from a cell culture dish, while providing a support, which is an essential condition of cell growth, by introducing the trypsin-free cell stamp system, and cells can be continuously supplied for a polymer-based fiber support without an additional subculture process since the empty space of a cell culture dish is filled as times passes. In addition, the artificial effects on cells can be minimized since the cells migrate to a polymer-based nano/micro-fiber support without other external stimulation, and thus the potency of stem cells is increased, thereby inducing more effective differentiation, such that the present invention, as a cell therapeutic agent, can be utilized in general fields of regenerative medicine and tissue engineering. 1. A trypsin-free cell stamp system.2. The system of claim 1 , wherein the stamp comprises a polymer-based nano/microfiber support.3. The system of claim 2 , wherein the support is porous for mechanical stability and cell culture.4. The system of claim 2 , wherein the support is used for supporting claim 2 , culturing claim 2 , or transplanting cells.5. The system of claim 2 , wherein the polymer is at least one selected from the group consisting of gelatin claim 2 , poly-alpha-ester group (poly-esters group) claim 2 , polyglycolic acid (PGA) claim 2 , polylactide (PLA) claim 2 , poly L-lactic acid (PLLA) claim 2 , poly D-lactic acid (PDLA) claim 2 , poly lactic-co-glycolic acid (PLGA) claim 2 , polycaprolactone (PCL) claim 2 , poly 2-hydroxyethyl methacrylate (pHEMA) claim 2 , polyethylene glycol (PEG) claim 2 , polypropylene glycol (PPG) claim 2 , polyhydroxybutyrate (PHB) which is polyhydroxyalkanoate claim 2 , polydioxanone (PDO claim 2 , PDS) claim 2 , polyurethane (PU) claim 2 , ...

Microfluidic model of the blood brain barrier

The invention relates to culturing brain endothelial cells, and optionally astrocytes and neurons in a fluidic device under conditions whereby the cells mimic the structure and function of the blood brain barrier. Culture of such cells in a microfluidic device, whether alone or in combination with other cells, drives maturation and/or differentiation further than existing systems.

MATERIALS AND METHODS FOR CELL CULTURE

Methods and compositions for improved cell growth and serum-reduced or -free cell culture media.

METHODS FOR TREATING RADIATION OR CHEMICAL INJURY

Methods for treating radiation or chemical injury are described that comprise administering to a subject a therapeutically effective amount of adherent stromal cells. Methods of preparing adherent stromal cells and pharmaceutical compositions comprising the cells are also described. 1126.-. (canceled)127. A method of mitigating reduction , resulting from exposure to radiation , in a subject's number of endogenous hematopoietic cells , comprising administering to said subject a composition comprising adherent stromal cells (ASC) , wherein the ASC are administered before said exposure , and wherein the ASC were cultured under three-dimensional culturing conditions.128. The method of claim 127 , wherein said ASC are administered by intramuscular injection.129. The method of claim 127 , wherein the origin of the ASC is placenta.130. The method of claim 129 , where at least 50% of the ASC are fetal cells.131. The method of claim 127 , wherein the origin of the ASC is selected from adipose tissue and bone marrow.132. The method of claim 127 , wherein said three-dimensional culturing conditions comprise culturing in a bioreactor.133. The method of claim 127 , wherein at least 70% of the ASC are positive for the marker CD200 claim 127 , as detected by flow cytometry compared to an isotype control.134. The method of claim 127 , wherein the ASC are administered 1 day before said exposure.135. A method of mitigating reduction claim 127 , resulting from exposure to a chemical claim 127 , in a subject's number of endogenous hematopoietic cells claim 127 , comprising administering to said subject a composition comprising adherent stromal cells (ASC) claim 127 , wherein the ASC are administered before said exposure claim 127 , and wherein the ASC were cultured under three-dimensional culturing conditions.136. The method of claim 135 , wherein said ASC are administered by intramuscular injection.137. The method of claim 135 , wherein the origin of the ASC is placenta.138. The ...

Engineered platform to generate 3d cardiac tissues

Described herein are a system, device, methods and compositions related to generating 3-dimensional cardiac tissues. Also described herein are a system, device, and methods of maturing 3-dimensional cardiac tissues and maintaining their viability in culture.

METHODS FOR MAINTAINING AND EXPANDING MESENCHYMAL STEM CELLS ON EXTRACELLULAR MATRIX COATED MICROCARRIERS

Disclosed are methods for coating microcarriers with a marrow stromal cell derived extracellular matrix, and maintaining and expanding mammalian mesenchymal stem cells on the marrow stromal cell derived extracellular matrix coated microcarriers in culture. 1. A method of maintaining and expanding mammalian mesenchymal stem cells in culture in an undifferentiated state , the method comprising: i. adding the microcarriers to a culture medium;', 'ii. adding mammalian bone marrow stromal cells to the culture medium;', 'iii. culturing the bone marrow stromal cells to produce the 3D extracellular matrix coating on the surface of the microcarriers;', 'iv. decellularizing the extracellular matrix coated microcarriers of the bone marrow stromal cells; and, 'a. producing a 3D extracellular matrix coating on the surface of microcarriers comprisingb. culturing the mammalian mesenchymal stem cells in the presence of the extracellular matrix coated microcarriers;wherein the extracellular matrix coating restrains differentiation of the mammalian mesenchymal stem cells.2. The method of claim 1 , wherein the extracellular matrix coating comprises collagen alpha-1 (XII) claim 1 , collagen alpha-3 (VI) claim 1 , EMILIN-1 claim 1 , serpin H1 claim 1 , thrombospondin-1 claim 1 , tenascin precursor (TN) (Human) claim 1 , transforming growth factor-beta-induced protein claim 1 , and vimentin.3. The method of claim 2 , wherein the extracellular matrix coating further comprises type I collagen claim 2 , type III collagen claim 2 , fibronectin claim 2 , decorin claim 2 , biglycan claim 2 , perlecan claim 2 , and laminin.4. The method of claim 3 , wherein the extracellular matrix coating further comprises at least one of syndecan-1 claim 3 , collagen type V claim 3 , or collagen type VI.56-. (canceled)7. The method of claim 1 , wherein the bone marrow stromal cells are isolated bone marrow mesenchymal stem cells.8. The method of claim 1 , wherein the mammalian mesenchymal stem cells are ...

In Vitro Pharmacokinetics/Pharmacodynamics Bellows Perfusion System for Enhancing Effectiveness of Cancer Chemotherapy

Provided herein is a continuous cell perfusion model system that provides useful pharmacokinetic and pharmacodynamic information on the application of new drugs or combinations of various agents in vitro to human cancer cell lines. Also provided are methods of using this system to individualize cancer treatment. 1. An in vitro cell culture system comprising:a) a compressible vessel comprising cell culture medium, a plurality of polymer flakes, and one or more porous membranes, wherein said polymer flakes have one or more cancer cells adhered thereto;b) a media vessel used to supply the cell culture medium to the compressible vessel, wherein the media vessel is attached to the compressible vessel via a first connection line having at least one access point where one or more anti-cancer drugs or chemotherapeutic agents can be added to the cell culture medium;c) a waste vessel used to remove waste products from the compressible vessel, wherein the waste vessel is attached to the compressible vessel via a second connection line having at least one access point where samples of the cell culture medium from the compressible vessel can be removed for analysis; andd) a hollow fiber tube that connects the media vessel and the compressible vessel.2. The in vitro cell culture system of claim 1 , wherein the hollow fiber tube provides an environment for endothelial cells to grow.3. The in vitro cell culture system of claim 2 , wherein the hollow fiber tube allows the study of the effect of anti-cancer drug or chemotherapeutic agent-induced anti-angiogenesis.4. The in vitro cell culture system of claim 2 , wherein the hollow fiber tube allows the study of the effect of angiogenesis induced by cancer cells grown in the compressible vessel in the control set and the study of anti-cancer drug or chemotherapeutic agent-induced anti-angiogenesis.5. The in vitro cell culture system of claim 1 , wherein the compressible vessel is a compressible (bellows) bottle.6. The in vitro cell ...

HOLLOW MICROCARRIER FOR SHEAR-FREE CULTURE OF ADHERENT CELLS IN BIOREACTORS

The present invention provides hollow microcarriers for cell culture. The hollow microcarriers form a shell around a hollow interior and can be opened to permit cell infiltration or harvesting. The hollow microcarriers protect cells from hydrodynamic shear stress without hindering the diffusion of nutrients in and out of their hollow interior. 1. A hollow microcarrier comprising a thin shell forming a three-dimensional structure having a hollow interior , the structure having a shape selected from the group consisting of: a sphere , an elongated sphere , a cylinder , a spheroid , and a polyhedron.2. The hollow microcarrier of claim 1 , wherein the shell comprises one or more holes claim 1 , gaps claim 1 , or apertures accessing the hollow interior.3. The hollow microcarrier of claim 1 , wherein the shell comprises a plurality of elongate leaflets claim 1 , each leaflet having a proximal end and a distal end claim 1 , wherein the plurality of leaflets are joined to each other at their proximal ends in a radial pattern claim 1 , and wherein the distal ends of the plurality of leaflets curl towards each other to form a substantially spherical shape having a hollow interior.4. The hollow microcarrier of claim 3 , comprising between 3 and 10 leaflets.5. The hollow microcarrier of claim 1 , wherein the shell comprises a plurality of elongate leaflets claim 1 , each leaflet having a proximal end and a distal end claim 1 , wherein the plurality of leaflets are joined to each other at their proximal ends in a first and a second radial pattern claim 1 , wherein the first and second radial patterns are joined to each other by the distal end of a leaflet claim 1 , and wherein the distal ends of the leaflets curl towards each other such that the first radial pattern and the second radial pattern each form a hemisphere of a substantially spherical shape having a hollow interior.6. The hollow microcarrier of claim 1 , wherein the shell comprises a plurality of elongate leaflets ...

Biomarker Detection Methods and Systems and Kits for Practicing Same

Aspects of the present disclosure include methods that include co-culturing a cell and a microparticle that includes a capture ligand, in a culture medium under conditions in which a biomarker produced by the cell is bound by the capture ligand. Such methods may further include detecting (e.g., by flow or mass cytometry) complexes that include the microparticle, the capture ligand, the biomarker, and a detection reagent. The methods may further include determining the proportion or number of cells among a heterogeneous cell population that produced the biomarker and/or the level of biomarker secreted by such cells. Compositions, systems and kits are also provided. 128.-. (canceled)29. A method , comprising: a heterogeneous cell population comprising a first subpopulation of cells that express a biomarker and a second subpopulation of cells; and', 'microparticles comprising capture ligands that specifically bind to the biomarker., 'co-culturing in a culture medium30. The method according to claim 29 , wherein the first subpopulation of cells secrete the biomarker.31. The method according to claim 30 , further comprising stimulating the first subpopulation of cells to secrete the biomarker claim 30 , wherein upon secretion of the biomarker claim 30 , the biomarker is bound by the capture ligands.32. The method according to claim 31 , wherein the stimulating comprises adding a stimulant to the culture medium.33. The method according to claim 32 , wherein the stimulant is selected from the group consisting of: an antibody claim 32 , an antigen claim 32 , a ligand claim 32 , a protein claim 32 , a lectin claim 32 , a nucleic acid claim 32 , a drug claim 32 , an allergen claim 32 , a peptide claim 32 , an interferon claim 32 , a chemokine claim 32 , an interleukin claim 32 , a lymphokine claim 32 , a tumor necrosis factor claim 32 , a CD molecule claim 32 , a cell claim 32 , a vaccine claim 32 , a parasite claim 32 , a fungus claim 32 , a sub-cellular component claim 32 , ...

Method for allogeneic cell therapy

A method of manipulating allogeneic cells for use in allogeneic cell therapy providing a composition of highly activated allogeneic T-cells which are infused into immunocompetent cancer patients to elicit a novel anti-tumor immune mechanism, or “Mirror Effect”. In contrast to current allogeneic cell therapy protocols where T-cells in the graft mediate the beneficial graft vs. tumor (GVT) and detrimental graft vs. host (GVH) effects, the allogeneic cells of the present invention stimulate host T-cells to mediate the “mirror” of these effects. The mirror of the GVT effect is the host vs. tumor (HVT) effect. The “mirror” of the GVH effect is the host vs. graft (HVG) effect The anti-tumor HVT effect occurs in conjunction with a non-toxic HVG rejection effect. The highly activated allogeneic cells of the invention can be used to stimulate host immunity in a complete HLA mis-matched setting in a patient. 1. A method for stimulating a coupled host vs. tumor and host vs. graft effect in a host which mirrors the coupled graft vs. tumor and graft vs. host effects of allogeneic transplant procedures comprising:selecting a composition of donor allogeneic cells; andadministering said allogeneic cells to the host who has not been pre-conditioned with immunosuppressive treatment.2. The method of wherein the composition of allogeneic cells include T-cells.3. The method of wherein the T-cells are predominately CD4+ T-cells.4. The method of wherein the CD4+ T-cells are predominantly Th1 cells.5. The method of wherein the T-cells are activated at the time of infusion.6. The method of wherein the T-cells are activated by cross-linking of CD3 and CD28 surface antigens. The present application is a divisional of and claims priority to U.S. patent application Ser. No. 15/041,642, filed Feb. 11, 2016, which is a divisional of and claims priority to U.S. patent application Ser. No. 14/173,494, filed Feb. 5, 2014 now U.S. Pat. No. 9,301,977, which is a divisional of and claims priority to U. ...

Metastasis and Adaptive Resistance Inhibiting Immunotherapy Combined Online Chemotherapy with Radiotherapy's tumor Seeking Extracellular Vesicles with siRNA and Chemotherapeutics

Mutated genome silencing with endogenous RNAi-siRNA and miRNA with near total cellular apheresis with pulse flow apheresis system and EV-exosome-RNA molecular apheresis with sucrose density gradient continuous flow ultracentrifugation combined with array centrifuge for both 50S higher and 50S lower proteomics and genomics apheresis and their fractionated purification with immobilized Tim4-Fc protein Ca2+ magnetic beads affinity chromatography (ACG) and immobilized metal ACG is disclosed. It purifies normal cell derived and tumor cell derived EVs-exosomes, proteomics and subcellular particles. Tumor-specific endogenous siRNA is generated from mutated RNA containing pre-miRNA hairpin through RNA-induced silencing complex (RISC) composed of Dicer, dsRNA binding protein TRBP, and AGO2. Incubating purified RSIC with pre-let-7 hairpin generates siRNA. SiRNA is bonded with T-EVs and T-cells to silence its evasion from tumor immunity. While on radiation therapy or surgery, a patient's blood is continuously processed with above systems. It delivers combined online radiotherapy, and tumor-seeking adoptive extracorporeal chemo-immunotherapy.

METHODS FOR TREATING RADIATION OR CHEMICAL INJURY

Methods for treating radiation or chemical injury are described that comprise administering to a subject a therapeutically effective amount of adherent stromal cells. Methods of preparing adherent stromal cells and pharmaceutical compositions comprising the cells are also described. 1. A method for treating a subject with a compromised endogenous hematopoietic system , comprising intramuscularly administering to the subject a pharmaceutical composition comprising a therapeutically effective amount of placenta-derived adherent stromal cells to induce repopulation of endogenous hematopoietic cells.2. The method of claim 1 , wherein exogenous hematopoietic stem cells are not administered to the subject.3. The method of claim 1 , wherein the subject has been exposed to radiation.4. The method of claim 1 , wherein the subject has been exposed to chemotherapy.5. The method of claim 1 , wherein the placental-derived adherent stromal cells are viable following expansion on three-dimensional carriers under conditions supporting cell expansion.6. The method of claim 5 , wherein the placental-derived adherent stromal cells are viable following detachment from said three-dimensional carriers into a pharmaceutical suspension.7. The method of claim 6 , wherein the placental-derived adherent stromal cells are viable following cryopreservation of said pharmaceutical suspension.8. The method of claim 5 , wherein the placental-derived adherent stromal cells exhibit enhanced immunosuppressive activity claim 5 , relative to placental-derived adherent stromal cells expanded on a two-dimensional substrate.9. The method of claim 5 , wherein the placental-derived adherent stromal cells exhibit enhanced secretion of Flt-3 ligand claim 5 , relative to placental-derived adherent stromal cells expanded on a two-dimensional substrate.10. The method of claim 5 , wherein the placental-derived adherent stromal cells exhibit enhanced secretion of IL-6 claim 5 , relative to placental-derived ...