CLEANING OF OF WILL FIRE FACTOR BY CATION EXCHANGER CHROMATOGRAPHY

POLYDISPERSE NATIVE ONES PSEUDOMONAS FLAGELLA (H) ANTIGENS (FAG) AND PROCEDURE FOR YOUR PRODUCTION.

Method of Isolation and Purification of Trypsin from Pronase and Use Thereof

Method for Producing Mature VWF From VWF Pro-peptide

The present invention relates to a method for producing a mature von Willebrand Factor (VWF) from von Willebrand Factor pro-peptide comprising the steps: - immobilizing VWF pro-peptide on an ion exchange resin, incubating the immobilized VWF pro-peptide with furin to obtain immobilized mature VWF, and - isolating mature VWF from the ion exchange resin by elution.

Pegylation of recombinant blood coagulation factors in the presence of bound antibodies

Protein purification by anion exchange chromatography

The present invention relates to a two-step method for the purification of divalent cation binding proteins with high yield and high purity on anion exchange resin materials, to divalent cation binding proteins obtainable by said method, and to a kit comprising means for carrying out said method.

METHOD OF PURIFICATION OF HYDROPHOBIC PROTEINS

The present invention relates to a method for obtaining highly purified hydrophobic proteins from cells by extraction using a buffer containing a detergent and removal of said detergent by hydroxyapatite (HA) column chromatography.

VIRUS FILTRATION OF CELL CULTURE MEDIA

The invention relates to a method for removing a viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. The method comprises subjecting said preparation to filtration for at least about 24 hours through a virus filter having an effective pore size of maximum about 75 nm. Further, the invention relates to the use of a virus filter in filtration of at least about 24 hours, wherein the virus filter has an effective pore size of maximum about 75 nm for the removal of viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. In some embodiments the filtration according to the invention operates at a volumetric capacity of at least about 2000 L/m2. Further, the invention relates to the use of a preparation, being a cell culture medium or at least a component of a cell culture medium obtainable according to method of the invention for cell culture; pharmaceutical, diagnostic and ...

METHOD FOR THE SEPARATION OF RECOMBINANT PRO-FACTOR IX FROM RECOMBINANT FACTOR IX

A method for separation of recombinant pro-Factor IX from recombinant Factor IX by a chromatographic method is described. According to a preferred embodiment, the mixture of both proteins is bound to an ion exchanger and pro-Factor IX and Factor IX are eluted separately from each other by buffer solutions with different salt concentrations and/or pH values. According to the method of the invention, pro-Factor IX and Factor IX are obtainable in highly pure form and free from the other factor respectively.

A METHOD OF PREPARING AND RECOVERING PROTEINS

There is disclosed the preparation and recovery of proteins, in particular of enzymes, by controlled proteolytic cleavage of pro-proteins, in particular pro-enzymes, in a single method step, wherein a proprotein-containing solution is contactd with a protease and with a solid carrier which has a higher affinity to the protein than to the pro-protein or to the funtionally inactive degradation products thereof, the pro-protein being proteolytically cleaved to said protein, and the protein being selectively separated by adsorption on the solid carrier.

SUBSTANTIALLY ANIMAL PROTEIN-FREE RECOMBINANT FURIN AND METHODS FOR PRODUCING SAME

SUBSTANTIALLY ANIMAL PROTEIN-FREE RECOMBINANT FURIN AND METHODS FOR PRODUCING SAMEAbstractThe present application relates to recombinant furin (rFurin) and methods for producing rFurin. More specifically, the invention relates to substantially animal protein-free rFurin and methods for producing substantially animal protein-free rFurin.No suitable figure ...

PROCEDURE FOR AUFREINIGUNG HYDROPHOBER PROTEINS

Method for purifying factor VIII/VWF complex by cation-exchange chromatography

Purification of VWF for increased removal of non-lipid enveloped viruses

The present invention provides methods for purifying Von Willebrand factor (VWF) for increased removal of non-lipid enveloped viruses.

Pegylation of recombinant blood coagulation factors in the presence of bound antibodies

The present invention relates to a proteinaceous construct comprising a blood coagulation factor, e.g., Factor VIII (FVIII), being bound to at least one water soluble polymer, including a poly(alkylene oxide) such as polyethylene glycol (PEG). Further the present invention relates to methods of preparing PEGylated blood coagulation factor, e.g., FVIII, in the presence of bound antibodies. The invention also relates to methods for prolonging the in vivo-half-life of blood coagulation factor, e.g., FVIII, in the blood of a mammal having a bleeding disorder associated with functional defects or deficiencies of blood coagulation factor, e.g., FVIII.

Method for the Isolation and Purification of Vitamin K-Dependent Proteins

A method for the separation of vitamin K-dependent proteins from non-vitamin K-dependent accompanying proteins is described wherein the method is characterized in that at least anion exchange chromatography and optionally affinity chromatography is carried out as well. The method is suitable especially for the purification of Factor II, VII, IX, X as well as Protein S, Protein C and Protein Z. With the aid of the method according to the invention a vitamin K-dependent protein is obtained which is present at a purity of 95%.

STABLE FACTOR VIII/ VON WILLEBRAND FACTOR COMPLEX

The invention relates to a stable factor VIII/ von Willebrand factor complex which contains high-molecular von Willebrand factor (vWF) multimers and is free of low-molecular vWF molecules and proteolytic vWF decomposition products. The invention also relates to a process for preparing said complex.

PURIFICATION OF VON WILLEBRAND FACTOR BY CATION EXCHANGE CHROMATOGRAPHY

Disclosed are a method of recovering vWF in which vWF at a low salt concentration is bound to a cation exchanger and vWF having a high specific activity is recovered by fractionated elution, as well as a preparation having purified vWF obtainable by this method.

Hücrelerin proteinsiz veya serumsuz yerleştirilmesi için ortam

Method of isolation and purification of trypsin from pronase protease and use thereof

The present invention is directed to methods of isolation and purification of Streptomyces griseus trypsin (SGT) from PRONASE protease in a single affinity chromatography step and uses of the purified SGT.

MEDIUM FOR PROTEIN AND SERUM-FREE THE CELL CULTURE

PROCEDURE FOR THE CHROMATOGRAPHI CLEANING AND/OR CRACKING OF OF WILL FIRE FACTOR FROM A VWF HÄLTIGEN RAW MATERIAL

PROCEDURE FOR THE DETERMINATION OF A KOLLAGEN BINDING SUBSTANCE, IN PARTICULAR THE ACTIVITY OF AN ADHESION PROTEIN

PROTHROMBIN-DERIVATE

Device for the quantification of prothrombin derivatives

PROCEDURE FOR THE ISOLATION AND CLEANING OF TRYPSIN FROM PRO NOSE AND YOUR USE

Method of concentrating shear-sensitive biopolymers using hollow fibre membranes

Protein purification by anion exchange chromatography

OLIGOPEPTIDE-FREE CELL CULTURE MEDIA

The present invention relates to oligopeptide-free cell culture media comprising at least 0.5 mg/L of a polyamine and to methods for cultivating cells in said oligopeptide-free cell culture media comprising at least 0.5 mg/L of a polyamine. The invention also relates to methods for expressing at least one protein in a medium comprising at least 0.5 mg/L of a polyamine an d to methods for producing at least one virus in a medium comprising at least 0.5 mg/L of a polyamine.

FORMULATION OF SUGAR SOLUTIONS FOR CONTINUOUS ULTRACENTRIFUGATION FOR VIRUS PURIFICATION

The present invention provides a method for purification of a virus or virus antigen comprising providing a virus preparation and centrifugation of said virus preparation in a gradient of a sugar established by the addition of two or more buffered sugar layers of different concentration. The method leads to higher yields and reduces unwanted aggregation of the virus or virus antigen by increasing the volume of the peak pool.

METHOD FOR PURIFYING RECOMBINANT ADAMTS13 AND OTHER PROTEINS AND COMPOSITIONS THEREOF

Provided herein are methods for purifying recombinant A Disintegrin-Iike and Metallopeptidase with Thrombospondin Type 1 Motif 13 (ADAMTS13) protein from a sample. The method comprises enriching for ADAMTS13 protein by chromatographically contacting the sample with hydroxyapatite under conditions that allow ADAMTS13 protein to appear in the eluate or supernatant from the hydroxylapatite. The methods may further comprise tandem chromatography with a mixed mode cation exchange/hydrophobic interaction resin that binds ADAMTS13 protein. Additional optional steps involve ultrafiltration/diafiltration, anion exchange chromatography, cation exchange chromatography, and viral inactivation. Also provided herein are methods for inactivating virus contaminants in protein samples, where the protein is immobilized on a support. Also provided herein are compositions of ADAMTS13 prepared according to said methods.

PURIFICATION OF VWF FOR INCREASED REMOVAL OF NON-LIPID ENVELOPED VIRUSES

The present invention provides methods for purifying Von Willebrand factor (VWF) for increased removal of non-lipid enveloped viruses.

PROCEDURE FOR THE PRODUCTION OF HIGHLY PURE VWF OR FACTOR VIII/VWF COMPLEX

PROCEDURE FOR THE PRODUCTION OF HIGHLY PURE ONE OF WILL FIRE FACTOR

STABLE FACTOR VIII/VWF COMPLEX

PROCEDURE FOR THE CLEANING OF FACTOR VIII/VWF COMPLEX OF MEANS CATION EXCHANGER CHROMATOGRAPHY

Medium for the protein-free and serum-free cultivation of cells

Method of purifying hepatitis A virus particles and vaccine preparation

Purification of VWF for Increased Removal of Non-Lipid Enveloped Viruses

The present invention provides methods for purifying Von Willebrand factor (VMF) for increased removal of non-lipid enveloped viruses.

Virus filtration of cell culture media

The invention relates to a method for removing a viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. The method comprises subjecting said preparation to filtration for at least about 24 hours through a virus filter having an effective pore size of maximum about 75 nm. Further, the invention relates to the use of a virus filter in filtration of at least about 24 hours, wherein the virus filter has an effective pore size of maximum about 75 nm for the removal of viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. In some embodiments the filtration according to the invention operates at a volumetric capacity of at least about 2000 L/m2. Further, the invention relates to the use of a preparation, being a cell culture medium or at least a component of a cell culture medium obtainable according to method of the invention for cell culture; pharmaceutical, diagnostic and ...

VIRUS FILTRATION OF CELL CULTURE MEDIA

The invention relates to a method for removing a viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. The method comprises subjecting said preparation to filtration for at least about 24 hours through a virus filter having an effective pore size of maximum about 75 nm. Further, the invention relates to the use of a virus filter in filtration of at least about 24 hours, wherein the virus filter has an effective pore size of maximum about 75 nm for the removal of viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. In some embodiments the filtration according to the invention operates at a volumetric capacity of at least about 2000 L/m 2. Further, the invention relates to the use of a preparation, being a cell culture medium or at least a component of a cell culture medium obtainable according to method of the invention for cell culture; pharmaceutical, diagnostic ...

SUBSTANTIALLY ANIMAL PROTEIN-FREE RECOMBINANT FURIN AND METHODS FOR PRODUCING SAME

The present invention relates to recombinant furin (rFurin) and methods for producing rFurin. More specifically, the invention relates to substantially animal protein-free rFurin and methods for producing substantially animal protein-free rFurin.

METHOD FOR ISOLATION OF HIGHLY PURE VON WILLEBRAND FACTOR

The invention relates to a method for isolation of highly pure von Willebrand Factor in which recombinant von Willebrand Factor (rvWF) is chromatographically purified by anion exchange chromatography on an anion exchanger of the quaternary amino type in a buffer solution comprising buffer substances and optionally salt. The buffer solutions are preferably free of stabilizers, amino acids and other additives. According to this method, highly pure recombinant vWF can be obtained, which is free from blood plasma proteins, especially free from Factor VIII, and is physiologically active. Further, the invention relates to a pharmaceutical preparation that contains rvtWF, which is comprised of multimers with a high structural integrity ...

Method for producing mature VWF from VWF pro-peptide

Targeted angiogenesis

Purification of von willebrand factor by cation exhcanger chromatography

METHODS OF PURIFYING RECOMBINANT ADAMTS13 AND OTHER PROTEINS AND COMPOSITIONS THEREOF

Provided herein are methods for purifying recombinant A Disintegrin-Iike and Metallopeptidase with Thrombospondin Type 1 Motif 13 (ADAMTS13) protein from a sample. The method comprises enriching for ADAMTS13 protein by chromatographically contacting the sample with hydroxyapatite under conditions that allow ADAMTS13 protein to appear in the eluate or supernatant from the hydroxylapatite. The methods may further comprise tandem chromatography with a mixed mode cation exchange/hydrophobic interaction resin that binds ADAMTS13 protein. Additional optional steps involve ultrafiltration/diafiltration, anion exchange chromatography, cation exchange chromatography, and viral inactivation. Also provided herein are methods for inactivating virus contaminants in protein samples, where the protein is immobilized on a support. Also provided herein are compositions of ADAMTS13 prepared according to said methods.

STABLE FACTOR VIII/VWF-COMPLEX

The invention relates to a stable factor VIII/ von Willebrand factor complex which contains high-molecular von Willebrand factor (vWF) multimers and is free of low-molecular vWF molecules and proteolytic v WF decomposition products. The invention also relates to a process for preparing said complex.

Method of production of purified hepatitis A virus particles and vaccine preparation

VIRUS FILTRATION OF CELL CULTURE MEDIA

PURIFICATION METHOD FOR DIVALENT CATION BINDING PROTEINS ON ANION EXCHANGE RESIN

The present invention relates to a method for the purification of divalent cation binding proteins with high purity on an anion exchange resin material, to divalent cation binding proteins obtainable by said method, and to a kit comprising means for carrying out said method. 1. A method for the purification of a divalent cation binding protein comprising the steps of:(a) loading an anion exchange resin material with the divalent cation binding protein in a loading buffer in the absence of divalent cations; and(b) eluting the divalent cation binding protein with an eluant comprising at least one divalent cation to form an eluate containing the divalent cation binding protein;wherein the eluant in step (b) has a pH higher than the pH of the loading buffer in step (a).2. The method of claim 1 , wherein the method further comprises the step of washing the loaded anion exchange resin material prepared in step (a) with a washing buffer in the absence of divalent cations claim 1 , prior to step (b) claim 1 , wherein the eluant has a higher pH than the pH of the washing buffer.3. The method according to claim 2 , wherein the pH of the eluant in step (b) is at least 0.5 pH units higher than the pH of the washing buffer.4. The method according to claim 2 , wherein the washing buffer has a conductivity that is equal or lower than the conductivity of the eluant in step (b).5. The method according to claim 2 , wherein the at least one divalent cation in step (b) is selected from the group consisting of Ca claim 2 , Be claim 2 , Ba claim 2 , Mg claim 2 , Mn claim 2 , Sr claim 2 , Zn claim 2 , Co claim 2 , Ni claim 2 , and Cu claim 2 , and a combination thereof.6. The method according to claim 2 , wherein the anion exchange resin material has a positively charged group which is selected from the group claim 2 , consisting of diethylaminoethane (DEAE) claim 2 , dimethylaminoethane (DMAE) claim 2 , trimethylaminoethyl (TMAE) claim 2 , polyethyleneimine (PEI) claim 2 , quaternary ...

VERFAHREN ZUR GEWINNUNG VON HOCHREINEM vWF ODER FACTOR VIII/vWF-KOMPLEX

Verfahren zur Trennung von vWF in hochmolekularen vWF und niedermolekularen vWF

Method of isolation and purification of trypsin from pronase and use thereof

Method of Isolation and Purification of Trypsin from Pronase and Use Thereof

Method of purification of hydrophobic proteins

The present invention relates to a method for obtaining highly purified hydrophobic proteins from cells by extraction using a buffer containing a detergent and removal of said detergent by hydroxyapatite (HA) column chromatography.

Prothrombin derivatives

PROTHROMBIN DERIVATIVES

The invention relates to novel prothrombin mutants or derivatives thereof having one or more changes in the protein sequence as against the natural protein, are either inactive or have an activity of 10 % at the most and preferably no more than 0.25 % of the natural protein and have a bonding capacity to natural ligands (natural or artificial anticoagulants) essentially corresponding to that of the natural protein. The description also relates to the use of mutated prothrombin mutants or derivatives as pharmaceutical preparations.

HIGH MOLECULAR AND LOW MOLECULAR FRACTIONS OF VON WILLEBRANDFACTOR

The present invention relates to a method for seperation of vWF into high molecular vWF and low molecular vWF which is characterized in that vWF is bound to an affinity support and is then eluted at different salt concentrations.

A METHOD OF RECOVERING HIGHLY PURIFIED VWF OR FACTOR VIII/VWF-COMPLEX

Disclosed is a method of recovering highly purified vWF or factor VIII/vWF-complex by means of immunoaffinity chromatography, which is characterized in that vWF or factor VIII/vWF-complex bound to an immunoadsorbent is eluted by means or an eluting agent containing as an essential active ingredient a zwitterion, while retaining the molecular integrity of vWF or of the factor VIII/vWF-complex. Furthermore, the invention relates to a preparation containing highly purified vWF having a specific activity of at least 100 U/mg protein, obtainable by immunoaffinity chromatography, as well as to the use of this preparation for producing a medicament for the treatment of vWD or of phenotypic hemophilia A.

A METHOD OF PURIFYING FACTOR VIII/VWF-COMPLEX BY MEANS OF CATION EXCHANGE CHROMATOGRAPHY

There is disclosed a method of recovering factor VIII/vWF-complex which is characterized in that factor VIII/vWF-complex from a protein solution is bound to a cation exchanger and is recovered by step-wise elution of factor VIII/vWF-complex, which particularly contains high-molecular vWF multimers, as well as a factor VIII/vWF-complex obtainable by means of cation exchange chromatography.

Method for purifying factor viii/vwf complex by cation-exchange chromatography

Purification of von willebrand factor by cation exhcanger chromatography

Method for producing mature VWF from VWF pro-peptide

Method for purifying recombinant ADAMTS13 and other proteins and compositions thereof

Provided herein are methods for purifying recombinant A Disintegrin-Iike and Metallopeptidase with Thrombospondin Type 1 Motif 13 (ADAMTS13) protein from a sample. The method comprises enriching for ADAMTS13 protein by chromatographically contacting the sample with hydroxyapatite under conditions that allow ADAMTS13 protein to appear in the eluate or supernatant from the hydroxylapatite. The methods may further comprise tandem chromatography with a mixed mode cation exchange/hydrophobic interaction resin that binds ADAMTS13 protein. Additional optional steps involve ultrafiltration/diafiltration, anion exchange chromatography, cation exchange chromatography, and viral inactivation. Also provided herein are methods for inactivating virus contaminants in protein samples, where the protein is immobilized on a support. Also provided herein are compositions of ADAMTS13 prepared according to said methods.

VIRUS FILTRATION OF CELL CULTURE MEDIA

The invention relates to a method for removing a viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. The method comprises subjecting said preparation to filtration for at least about 24 hours through a virus filter having an effective pore size of maximum about 75 nm. Further, the invention relates to the use of a virus filter in filtration of at least about 24 hours, wherein the virus filter has an effective pore size of maximum about 75 nm for the removal of viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. In some embodiments the filtration according to the invention operates at a volumetric capacity of at least about 2000 L/m 2. Further, the invention relates to the use of a preparation, being a cell culture medium or at least a component of a cell culture medium obtainable according to method of the invention for cell culture; pharmaceutical, diagnostic and/or cosmetic preparations as well as in food preparations.

METHOD FOR ISOLATION OF HIGHLY PURE VON WILLEBRAND FACTOR

The invention concerns a process for extracting high-purity von Willebrand factor wherein recombinant von Willebrand factor (rvWF) is chromatographically purified by anion exchange chromatography on a quaternary amino-type anion exchanger in a buffer solution comprising buffer substances and optionally salt. The buffer solutions are preferably free from stabilizers, amino acids and other additives. This process produces high-purity recombinant vWF which is free from blood plasma proteins, in particular factor VIII, and is physiologically active. The invention further concerns a pharmaceutical preparation which contains rvWF and is composed of multimers having high structural integrity.

A METHOD OF PURIFYING FACTOR VIII/VWF-COMPLEX BY MEANS OF CATION EXCHANGE CHROMATOGRAPHY

The invention relates to a method for obtaining factor VIII/vWF complex, characterized in that factor VIII/vWF complex from a protein solution is bonded with a cation-exchanger and factor VIII/vWF complex especially containing vWF multimers of high molecular weight is obtained by gradual elution. The invention also relates to a factor VIII/vWF complex which can be obtained by cation-exchange chromatography.

METHODS OF CONCENTRATING SHEAR-SENSITIVE BIOPOLYMERS

The invention relates generally to methods of concentrating mixtures including shear sensitive biopolymers, such as von Willebrand Factor. Conventional methods of concentrating biopolymers impart too much shear stress, which causes the degradation of shear sensitive biopolymers. The methods disclosed herein reduce the shear stress while maintaining a high rate of filtrate flux. Disclosed herein is a method for concentrating shear sensitive biopolymers including flowing a mixture with a shear sensitive biopolymer into a hollow fiber dialysis module to form a retentate having a shear sensitive biopolymer concentration that is greater than that of the mixture. Hollow fiber dialysis modules have high filtrate fluxes and low shear rates at low flow rates. This ensures a high product yield and minimal loss of shear sensitive biopolymers.

METHOD FOR ISOLATING AND PURIFYING TRYPSIN FROM PRONASE, AND USE OF THE SAME

PROBLEM TO BE SOLVED: To provide a method for isolating and purifying Streptomyces griseus trypsin (SGT) from a pronase in a single affinity chromatography process, and a method for using the purified SGT. SOLUTION: This method for isolating Streptomyces griseus trypsin (SGT) from Streptomyces griseus pronase comprises a process of bringing the pronase in contact with an immobilized affinity part selected from the group consisting of amidine, guanadine or an amine-containing species, and a process of selectively eluting the SGT from the immobilized affinity part by using an eluting liquid containing an eluent selected from the amidine, guanadine or amine-containing species. COPYRIGHT: (C)2011,JPO&INPIT ...

Method for obtaining high-purity VWF or factor VIIU/VWF complex

Targeted angiogenesis

Substantially animal protein-free recombinant furin and methods for producing same

Method of purification of hydrophobic proteins

METHOD FOR PRODUCING MATURE VWF FROM VWF PRO-PEPTIDE

The present invention relates to a method for producing a mature von Willebrand Factor (VWF) from von Willebrand Factor pro-peptide comprising the steps: - immobilizing VWF pro-peptide on an ion exchange resin, incubating the immobilized VWF pro-peptide with furin to obtain immobilized mature VWF, and - isolating mature VWF from the ion exchange resin by elution.

PEGYLATION OF RECOMBINANT BLOOD COAGULATION FACTORS IN THE PRESENCE OF BOUND ANTIBODIES

The present invention relates to a proteinaceous construct comprising a blood coagulation factor, e.g., Factor VIII (FVIII), being bound to at least one water soluble polymer, including a poly(alkylene oxide) such as polyethylene glycol (PEG). Further the present invention relates to methods of preparing PEGylated blood coagulation factor, e.g., FVIII, in the presence of bound antibodies. The invention also relates to methods for prolonging the in vivo-half-life of blood coagulation factor, e.g., FVI-II, in the blood of a mammal having a bleeding disorder associated with functional defects or deficiencies of blood coagulation factor, e.g., FVIII.

PEGYLATION OF RECOMBINANT BLOOD COAGULATION FACTORS IN THE PRESENCE OF BOUND ANTIBODIES

The present invention relates to a proteinaceous construct comprising a blood coagulation factor, e.g., Factor VIII (FVIII), being bound to at least one water soluble polymer, including a poly(alkylene oxide) such as polyethylene glycol (PEG). Further the present invention relates to methods of preparing PEGylated blood coagulation factor, e.g., FVIII, in the presence of bound antibodies. The invention also relates to methods for prolonging the in vivo-half-life of blood coagulation factor, e.g., FVIII, in the blood of a mammal having a bleeding disorder associated with functional defects or deficiencies of blood coagulation factor, e.g., FVIII.

METHOD OF CONCENTRATING SHEAR-SENSITIVE BIOPOLYMERS USING HOLLOW FIBRE MEMBRANES

The invention relates generally to methods of concentrating mixtures including shear sensitive biopolymers, such as von Willebrand Factor. Conventional methods of concentrating biopolymers impart too much shear stress, which causes the degradation of shear sensitive biopolymers. The methods disclosed herein reduce the shear stress while maintaining a high rate of filtrate flux. Disclosed herein is a method for concentrating shear sensitive biopolymers including flowing a mixture with a shear sensitive biopolymer into a hollow fiber dialysis module to form a retentate having a shear sensitive biopolymer concentration that is greater than that of the mixture. Hollow fiber dialysis modules have high filtrate fluxes and low shear rates at low flow rates. This ensures a high product yield and minimal loss of shear sensitive biopolymers.

HIGH MOLECULAR AND LOW MOLECULAR FRACTIONS OF VON WILLEBRAND FACTOR

The present invention relates to a method for seperation of vWF into high molecular vWF and low molecular vWF which is characterized in that vWF is bound to an affinity support and is then eluted at different salt concentrations.

PURIFICATION OF VON WILLEBRAND FACTOR BY CATION EXCHANGER CHROMATOGRAPHY

The invention relates to a method for obtaining vWF, whereby vWf is bonded to a cation exchanger at a low salt concentration and vWf with high specific activity is obtained by fractional elution. The invention also relates to a preparation with purified vWf which can be obtained by this method.

A METHOD OF CHROMATOGRAPHICALLY PURIFYING OR FRACTIONATING, RESPECTIVELY, VON WILLEBRAND FACTOR FROM A VWF-CONTAINING STARTING MATERIAL

The invention concerns a method of chromatographically purifying or fractionating von Willebrand factor (vWF) from a vWF-containing starter material, the method comprising the following steps: adsorption of the vWF from the starter material on avid collagen immobilized on a carrier; separation of the non-absorbed part and optionally washing of the carrier; elution of the vWF from the immobilized collagen; and extraction of the purified vWF. The invention also concerns a pharmaceutical preparation comprising biologically active vWF which is stably bonded to collagen.

Formulation of Sugar Solutions for Continuous Ultracentrifugation for Virus Purification

The present invention provides a method for purification of a virus or virus antigen comprising providing a virus preparation and centrifugation of said virus preparation in a gradient of a sugar established by the addition of two or more buffered sugar layers of different concentration. The method leads to higher yields and reduces unwanted aggregation of the virus or virus antigen by increasing the volume of the peak pool.

Prothrombin derivatives

Purification method for vitamin K dependent proteins by anion exchange chromatography

The present invention relates to a method for the purification of Vitamin K dependent proteins with high yield and high purity, particularly enriched in active protein, on anion exchange resin materials, to Vitamin K dependent proteins obtainable by said method, and to a kit comprising means for carrying out said method.

Targeted angiogenesis

Oligopeptide-free cell culture media

METHOD OF CONCENTRATING SHEAR-SENSITIVE BIOPOLYMERS USING HOLLOW FIBRE MEMBRANES

The invention relates generally to methods of concentrating mixtures including shear sensitive biopolymers, such as von Willebrand Factor. Conventional methods of concentrating biopolymers impart too much shear stress, which causes the degradation of shear sensitive biopolymers. The methods disclosed herein reduce the shear stress while maintaining a high rate of filtrate flux. Disclosed herein is a method for concentrating shear sensitive biopolymers including flowing a mixture with a shear sensitive biopolymer into a hollow fiber dialysis module to form a retentate having a shear sensitive biopolymer concentration that is greater than that of the mixture. Hollow fiber dialysis modules have high filtrate fluxes and low shear rates at low flow rates. This ensures a high product yield and minimal loss of shear sensitive biopolymers.

PURIFICATION METHOD FOR DIVALENT CATION BINDING PROTEINS ON ANION EXCHANGE RESIN

The present invention relates to a method for the purification of divalent cation binding proteins with high purity on an anion exchange resin material, to divalent cation binding proteins obtainable by said method, and to a kit comprising means for carrying out said method.

A METHOD OF PREPARING AND RECOVERING PROTEINS

There is disclosed the preparation and recovery of proteins, in particular of enzymes, by controlled proteolytic cleavage of pro-proteins, in particular pro-enzymes, in a single method step, wherein a pro--protein-containing solution is contactd with a protease and with a solid carrier which has a higher affinity to the protein than to the pro-protein or to the funtionally inactive degradation products thereof, the pro-protein being proteolytically cleaved to said protein, and the protein being selectively separated by adsorption on the solid carrier.

METHOD FOR THE ISOLATION AND PURIFICATION OF VITAMIN K-DEPENDENT PROTEINS

A method for the separation of vitamin K-dependent proteins from non-vitamin K-dependent accompanying proteins is described wherein the method is characterized in that at least anion exchange chromatography and optionally affinity chromatography is carried out as well. The method is suitable especially for the purification of Factor II, VII, IX, X as well as Protein S, Protein C and Protein Z. With the aid of the method according to the invention a vitamin K-dependent protein is obtained which is present at a purity of 95%.

A METHOD OF RECOVERING HIGHLY PURIFIED VWF OR FACTOR VIII/VWF-COMPLEX

The invention relates to a method for obtaining high-purity vWF or factor VIII/vWF complex by immunoaffinity chromatography, characterized in that a vWF or factor VIII/vWF complex bound with an immune adsorbing agent is eluted with an elution agent containing a zwitterion as an essential active part, whilst preserving the molecular integrity of the vWF or factor VIII/vWF complex. The invention also relates to a preparation containing high-purity vWF with a specific activity of at least 100 U/mg protein obtainable by immunoaffinity chromatography and to the use of this preparation for producing a medicament for treating vWD or phenotypic hemophilia A.

Virus filtration of cell culture media

The invention relates to a method for removing a viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. The method comprises subjecting said preparation to filtration for at least about 24 hours through a virus filter having an effective pore size of maximum about 75 nm. Further, the invention relates to the use of a virus filter in filtration of at least about 24 hours, wherein the virus filter has an effective pore size of maximum about 75 nm for the removal of viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. In some embodiments the filtration according to the invention operates at a volumetric capacity of at least about 2000 L/m2. Further, the invention relates to the use of a preparation, being a cell culture medium or at least a component of a cell culture medium obtainable according to method of the invention for cell culture; pharmaceutical, diagnostic ...

PURIFICATION OF VWF FOR INCREASED REMOVAL OF NON-LIPID ENVELOPED VIRUSES

PURIFICATION OF VWF FOR INCREASED REMOVAL OF NON LIPID ENVELOPED VIRUSES Abstract 5 The present invention provides methods for purifying Von Willebrand factor (VWF) for increased removal of non-lipid enveloped viruses. No suitable figure 32 ...

VERFAHREN ZUR GEWINNUNG VON HOCHREINEM VON WILLEBRAND-FAKTOR

Oligopeptide-free cell culture media

The present invention relates to oligopeptide-free cell culture media comprising at least 0.5 mg/L of a polyamine and to methods for cultivating cells in said oligopeptide-free cell culture media comprising at least 0.5 mg/L of a polyamine. The invention also relates to methods for expressing at least one protein in a medium comprising at least 0.5 mg/L of a polyamine and to methods for producing at least one virus in a medium comprising at least 0.5 mg/L of a polyamine.

METHOD OF PRODUCTION OF PURIFIED HEPATITIS A VIRUS PARTICLES AND VACCINE PREPARATION

Method for Purifying Recombinant ADAMTS13 and Other Proteins and Compositions Thereof

Abstract Provided herein are methods for purifying recombinant A Disintegrin-like and Metallopeptidase with Thrombospondin Type 1 Motif 13 (ADAMTS 13) protein from a sample. The method comprises enriching for ADAMTS 13 protein by chromatographically contacting the sample with hydroxyapatite under conditions that allow ADAMTS 13 protein to appear in the eluate or supernatant from the hydroxylapatite. The methods may further comprise tandem chromatography with a mixed mode f4 cation exchange/hydrophobic interaction resin that binds ADAMTS 13 protein. Additional optional steps involve ultrafiltration/diafiltration, anion exchange chromatography, cation exchange chromatography, and viral inactivation. Also provided herein are methods for inactivating virus contaminants in protein samples, where the protein is immobilized on a support. Also provided herein are compositions of ADAMTS 13 prepared according to said methods.

PROCESS FOR EXTRACTING AND PURIFYING RECOMBINANT, NON-LIPIDISED OSP-PROTEIN

A recombinant, non-lipidised OspC is disclosed, as well as a process for preparing Osp-proteins and vaccines which contain these recombinant proteins.

METHOD OF PURIFICATION OF HYDROPHOBIC PROTEINS

The present invention relates to a method for obtaining highly purified hydrophobic proteins from cells by extraction using a buffer containing a detergent and removal of said detergent by hydroxyapatite (HA) column chromatography.

FORMULATION OF SUGAR SOLUTIONS FOR CONTINUOUS ULTRACENTRIFUGATION FOR VIRUS PURIFICATION

The present invention provides a method for purification of a virus or virus antigen comprising providing a virus preparation and centrifugation of said virus preparation in a gradient of a sugar established by the addition of two or more buffered sugar layers of different concentration. The method leads to higher yields and reduces unwanted aggregation of the virus or virus antigen by increasing the volume of the peak pool.

METHOD OF ISOLATION AND PURIFICATION OF TRYPSIN FROM PRONASE PROTEASE AND USE THEREOF

The present invention provides methods of isolation and purification of Streptomyces griseus trypsin (SGT) from PRONASE protease mixture in a single affinity chromatography step and uses of the purified SGT. 1. A method of using a purified preparation of Streptomyces griseus trypsin (SGT) having a specific activity of at least about 25×10U/mg protein , and further comprising arginine , in a biological process , wherein the biological process selected from the group consisting of production of a virus or virus antigen , production of a recombinant product from recombinant cells , production of a cellular biomass grown in serum and protein free media , purification of a virus or virus antigen , and the activation of proteins from pro-proteins.2. The method of claim 1 , wherein the biological process is the production of virus or virus antigen claim 1 , the method comprising providing a cell culture of cells claim 1 , wherein the cells are being passaged and subcultured using the purified preparation of SGT claim 1 , growing the cells to a biomass claim 1 , infecting the cells of the biomass with a virus claim 1 , and incubating said cells to propagate said virus.3. The method of claim 2 , wherein the virus is selected from the group consisting of paramyxoviridae claim 2 , orthomyxoviridae claim 2 , and rotaviridae claim 2 , and the method further comprising adding the purified preparation to activate said virus and harvesting said virus produced.4. The method according to claim 3 , further comprising purifying said virus produced.5. A virus or virus antigen preparation obtained by the method of .6. The method of claim 1 , wherein the biological process is the production of a recombinant product from recombinant cells the method comprising the steps of providing cell culture biomass of recombinant cells claim 1 , wherein said cells have been passaged and subcultured using the purified preparation claim 1 , and culturing said cells under conditions claim 1 , whereby ...

METHOD FOR THE PURIFICATION OF RECOMBINANT BLOOD COAGULATION FACTOR IX ENRICHED IN SULFATED AND/OR PHOSPHORYLATED MOLECULES

The present invention relates to a method for the purification of rFIX using anion exchange chromatography in the pseudo-affinity mode, wherein said method comprises a wash step with a wash buffer having a salt concentration of more than 200 mM. The purification according to the invention provides a method to enrich rFIX molecules which have been posttranslationally modified by sulfation and/or phosphorylation. The present invention further relates to purified rFIX compositions enriched in monosulfated and/or monophosphorylated rFIX molecules. 17-. (canceled)8. A purified composition comprising rFIX obtained by a method , the method for obtaining the composition comprising the steps:a) loading a starting composition comprising rFIX onto an anion exchange column.,b) washing the anion exchange material using a wash buffer which has a salt concentration of more than 200 mM (High Salt Wash Buffer), andc) eluting the rFIX from the anion exchange material using an elution buffer comprising divalent cations, andd) collecting the eluate.9. The purified composition according to claim 8 , wherein said purified composition comprises an increased relative amount of monosulfated and/or monophosphorylated rFIX molecules compared to the relative amount of monosulfated and/or monophosphorylated rFIX present in the High Salt Wash Buffer fraction.10. The purified composition according to claim 8 , wherein said purified composition comprises a relative amount of monosulfated and monophosphorylated rFIX molecules which is increased at least 3 fold compared to the relative amount of monosulfated and monophosphorylated rFIX present in the High Salt Wash Buffer fraction.11. The purified composition according to claim 8 , wherein said purified composition comprises a relative amount of at least 30% of monosulfated and/or monophosphorylated rFIX molecules.12. The purified composition according to claim 8 , wherein said purified composition comprises a relative amount of at least 50% of ...

Virus Filtration of Cell Culture Media

The invention relates to a method for removing a viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. The method comprises subjecting said preparation to filtration for at least about 24 hours through a virus filter having an effective pore size of maximum about 75 nm. Further, the invention relates to the use of a virus filter in filtration of at least about 24 hours, wherein the virus filter has an effective pore size of maximum about 75 nm for the removal of viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. In some embodiments the filtration according to the invention operates at a volumetric capacity of at least about 2000 L/m. Further, the invention relates to the use of a preparation, being a cell culture medium or at least a component of a cell culture medium obtainable according to method of the invention for cell culture; pharmaceutical, diagnostic and/or cosmetic preparations as well as in food preparations. 1. A method for removing a viral contaminant from a preparation , said preparation being a cell culture medium or at least one component of a cell culture medium , comprising the step of:a) subjecting said preparation to filtration for at least about 24 hours through a virus filter having an effective pore size of maximum 75 nm.2. The method of claim 1 , wherein the filtration operates at a volumetric capacity of at least about 2000 L/m claim 1 , at least about 3000 L/m claim 1 , or at least about 5000 L/m.3. The method of claim 1 , wherein said preparation is subjected to filtration for at least about 48 hours up to about 7 months or at least about 72 hours up to about 3 months.4. The method of claim 1 , further comprising the step:b) feeding the filtrate to a cell culture medium or to at least one component of a cell culture medium.5. The method of or claim 1 , wherein the filtration is a continuous filtration.6. The method ...

MEDIUM FOR THE PROTEIN-FREE AND SERUM-FREE CULTIVATION OF CELLS

A medium is described for the protein-free and serum-free cultivation of cells, especially mammalian cells, whereby the medium contains a proportion of soy hydrolysate. 1. An animal protein-free and serum-free cell culture medium , the medium comprising a soy hydrolysate having a total nitrogen content of between 7.6% and 11.4% , and the medium comprising0.001-1 g/L L-asparagine,0.001-1 g/L L-cysteine,0.001-1 g/L L-cystine,0.001-1.5 g/L L-proline,0.001-1 g/L L-tryptophan, and0.05-1 g/L L-glutamine.2. The animal protein-free and serum-free cell culture medium of claim 1 , wherein the medium comprises an endotoxin content of <500 U/g.3. The animal protein-free and serum-free cell culture medium of claim 1 , wherein the medium comprises more than 10 wt. % ultrafiltered soy hydrolysate based on the total dry weight of the medium claim 1 , and wherein at least 40% of the soy hydrolysate has a molecular weight of ≦500 daltons.4. The animal protein-free and serum-free cell culture medium of claim 3 , wherein at least 50% of the soy hydrolysate has a molecular weight of ≦500 daltons.5. The animal protein-free and serum-free cell culture medium of claim 3 , wherein at least 55% of the soy hydrolysate has a molecular weight of ≦500 daltons.6. The animal protein-free and serum-free cell culture medium of claim 1 , wherein the medium contains ultrafiltered soy hydrolysate.7. The animal protein-free and serum-free cell culture medium of claim 1 , wherein the medium comprises an amino acid.8. The animal protein-free and serum-free cell culture medium of claim 7 , wherein the amino acid is selected from the group consisting of L-asparagine claim 7 , L-cysteine claim 7 , L-cystine claim 7 , L-proline claim 7 , L-tryptophan claim 7 , L-glutamine and mixtures thereof.9. The animal protein-free and serum-free cell culture medium of claim 1 , wherein the animal protein-free and serum-free cell culture medium further comprises: 1 to 100 g/L synthetic minimal medium; 0.05-1 g/L glutamine ...

PURIFICATION METHOD FOR VITAMIN K DEPENDENT PROTEINS BY ANION EXCHANGE CHROMATOGRAPHY

The present invention relates to a method for the purification of Vitamin K dependent proteins with high yield and high purity, particularly enriched in active protein, on anion exchange resin materials, to Vitamin K dependent proteins obtainable by said method, and to a kit comprising means for carrying out said method. 1. A method for the purification of a Vitamin K dependent protein comprising the steps of:(a) loading an anion exchange resin material (in the following also “the first anion exchange resin material”) with the Vitamin K dependent protein in a loading buffer in the absence or low concentration of divalent cations, optionally followed by one to three wash steps; 1-3 mM calcium is comprised in the eluant', 'the eluant has a conductivity of 14-23 mS/cm (25° C.), and', 'the pH of the eluant is between 7.0 and 9.0., '(b) eluting the Vitamin K dependent protein with an eluant comprising calcium and a counter-anion to form an eluate containing the Vitamin K dependent protein, wherein2. The method for purification of further comprising the following steps:(c) diluting the obtained eluate pool, ((1) to optionally lower the conductivity), and increasing the concentration of the calcium,(d) loading a second anion exchange resin material with the eluate as obtained after step (c); and(e) collecting the flow-through containing the Vitamin K dependent protein.3. The method according to or claim 1 , wherein in step (b):{'sup': '++', 'the conductivity is between 18 and 20 mS/cm (25° C.) at a pH of 7.0-7.4 and 1 mM Ca,'}{'sup': '++', 'the conductivity is between 18.5 and 21 mS/cm (25° C.) at a pH of 7.4-7.6 and 1 mM Ca,'}{'sup': '++', 'the conductivity is between 19 and 22.5 mS/cm (25° C.) at a pH of 7.6-8.0 and 1 mM Ca,'}{'sup': '++', 'the conductivity is between 20 and 23 mS/cm (25° C.) at a pH of 8.0-9.0 and 1 mM Ca,'}{'sup': '++', 'the conductivity is between 15 and 17.0 mS/cm (25° C.) at a pH of 7.0-7.4 and 2 mM Ca,'}{'sup': '++', 'the conductivity is between 16 ...

FORMULATION OF SUGAR SOLUTIONS FOR CONTINUOUS ULTRACENTRIFUGATION FOR VIRUS PURIFICATION

The present invention provides a method for purification of a virus or virus antigen comprising providing a virus preparation and centrifugation of said virus preparation in a gradient of a sugar established by the addition of two or more buffered sugar layers of different concentration. The method leads to higher yields and reduces unwanted aggregation of the virus or virus antigen by increasing the volume of the peak pool. 1. A method for purifying a virus comprising a buffered sugar solution having a density of 1.15 kg/l to 1.23 kg/l and comprising a first physiological buffer and', '(ii) a buffered sugar solution having a density of 1.23 kg/l to 1.32 kg/l and which is higher than the buffered sugar solution (i), and comprising a physiological buffer which is the same or different from the physiological buffer of (i),', 'wherein the volume ratio of buffered sugar solution (i) to buffered sugar solution (ii) is greater than or equal to 3:1;, 'providing'}providing a virus preparation;centrifuging the virus preparation and buffered sugar solutions (i) and (ii) in a ultracentrifuge comprising an ultracentrifuge rotor to obtain a peak pool; andextracting the peak pool to obtain the virus.2. The method of claim 1 , wherein the volume of buffered sugar solutions (i) and (ii) is between 5% to 100% of the volume of the ultracentrifugation rotor.3. The method of claim 1 , wherein the peak pool has a density between 1.20 kg/l to 1.24 kg/l.4. (canceled)5. The method of claim 1 , wherein one of said buffers has a concentration of between 5 mM to 50 mM.6. The method of claim 5 , wherein said buffer has a concentration of between 15 mM to 30 mM.7. The method of claim 6 , wherein said buffer has a concentration of between 18 mM to 25 mM.8. The method of claim 1 , wherein the step of centrifuging the virus preparation and buffered sugar solutions is performed with a relative centrifugation of at least 20 claim 1 ,000 g.9. The method of claim 8 , wherein said relative ...

OLIGOPEPTIDE-FREE CELL CULTURE MEDIA

The present invention relates to oligopeptide-free cell culture media comprising at least 0.5 mg/L of a polyamine and to methods for cultivating cells in said oligopeptide-free cell culture media comprising at least 0.5 mg/L of a polyamine. The invention also relates to methods for expressing at least one protein in a medium comprising at least 0.5 mg/L of a polyamine and to methods for producing at least one virus in a medium comprising at least 0.5 mg/L of a polyamine. 1. A method for expressing a coagulation factor VIII protein , comprising the steps of:(a) providing a culture of CHO cells;(b) introducing at least one nucleic acid sequence comprising a sequence coding for the coagulation factor VIII protein into the cells;(c) selecting the cells carrying the nucleic acid sequence; and(d) expressing the coagulation factor VIII protein in an oligopeptide-free chemically defined medium comprising DMEM:HAM's F12 (1:1) basal medium, putrescine at a concentration of at least 1 mg/L, and Fe(II) and Cu(II) at a level greater than the amount in basal DMEM:HAM's F12 (1:1), wherein the cells are cultivated by chemostat cultivation.2. The method of claim 1 , wherein the putrescine is present in the culture medium at a concentration ranging from 1 to 20 mg/L.3. The method of claim 1 , wherein the chemically defined medium is supplemented with L-glutamine claim 1 , ascorbic acid claim 1 , ethanolamine claim 1 , sodium selenite claim 1 , and a non-ionic surfactant. This application is a continuation of U.S. patent application Ser. No. 13/035,696 filed Feb. 25, 2011, which is a continuation of U.S. patent application Ser. No. 11/649,694 filed Jan. 3, 2007, which claims benefit of U.S. provisional application Ser. No. 60/756,419 filed Jan. 4, 2006, which applications are herein incorporated by reference.The present invention relates to oligopeptide-free cell culture media comprising at least 0.5 mg/L of a polyamine and to methods for cultivating cells in said oligopeptide-free ...

MEDIUM FOR THE PROTEIN-FREE AND SERUM-FREE CULTIVATION OF CELLS

A medium is described for the protein-free and serum-free cultivation of cells, especially mammalian cells, whereby the medium contains a proportion of soy hydrolysate. 1. A method of adapting recombinant cells to serum-free and animal protein-free medium , the method comprising:a. culturing recombinant cells in serum-containing medium,b. transferring said cells into a serum-free and animal protein-free medium, wherein the medium comprises soy hydrolysate wherein at least 40% of said soy hydrolysate has a molecular weight of ≦500 Daltons, andc. culturing said cells in said serum-free and animal protein-free medium.2. The method of claim 1 , wherein the medium contains a quantity in excess of 10 wt % soy hydrolysate based on the total dry weight of the medium.3. The method of claim 1 , wherein said medium contains ultrafiltered soy hydrolysate.4. The method of claim 3 , wherein the medium contains purified soy hydrolysate.5. The method of claim 1 , wherein the soy hydrolysate has an endotoxin content of <500 U/g.6. The method of claim 1 , wherein at least 500 of the soy hydrolysate has a molecular weight of ≦500 Daltons.7. The method of claim 6 , wherein at least 550 of the soy hydrolysate has a molecular weight of ≦500 Daltons.8. The method of claim 1 , wherein the medium also contains an amino acid.9. The method of claim 7 , wherein the amino acid is selected from the group consisting of L-asparagine claim 7 , L-cysteine claim 7 , L-cystine claim 7 , L-proline claim 7 , L-tryptophan claim 7 , and L-glutamine claim 7 , or mixtures thereof.10. The method of claim 1 , wherein the medium also contains one or more auxiliary substances selected from the group consisting of buffer substances claim 1 , oxidation stabilizers claim 1 , stabilizers to counteract mechanical stress claim 1 , and protease inhibitors.11. The method of claim 1 , wherein the cells contain a coding sequence for a recombinant blood factor claim 1 , and are capable of expressing the recombinant blood ...

METHODS OF PURIFYING RECOMBINANT ADAMTS13 AND OTHER PROTEINS AND COMPOSITIONS THEREOF

Provided herein are methods for purifying recombinant A Disintegrin-like and Metallopeptidase with Thrombospondin Type 1 Motif 13 (ADAMTS13) protein from a sample. The method comprises enriching for ADAMTS13 protein by chromatographically contacting the sample with hydroxyapatite under conditions that allow ADAMTS13 protein to appear in the eluate or supernatant from the hydroxylapatite. The methods may further comprise tandem chromatography with a mixed mode cation exchange/hydrophobic interaction resin that binds ADAMTS13 protein. Additional optional steps involve ultrafiltration/diafiltration, anion exchange chromatography, cation exchange chromatography, and viral inactivation. Also provided herein are methods for inactivating virus contaminants in protein samples, where the protein is immobilized on a support. Also provided herein are compositions of ADAMTS13 prepared according to said methods. 1. A method for purifying recombinant a disintegrin-like and metallopeptidase with thrombospondin type 1 motif 13 (ADAMTS13) protein from a sample comprising ADAMTS13 protein and non-ADAMTS13 impurities , the method comprising chromatographically contacting the sample with hydroxyapatite under conditions that allow said ADAMTS13 protein to appear in an eluate or a supernatant from said hydroxyapatite.2. The method according to claim 1 , further comprising chromatographically contacting said eluate with a cation exchange/hydrophobic interaction resin that binds said ADAMTS13 protein.3. The method according to or claim 1 , further comprising chromatographically contacting said sample with an anion exchange resin and eluting said ADAMTS13 protein from said anion exchange resin before chromatographic contact with said hydroxyapatite.4. The method according to or claim 1 , further comprising concentrating said ADAMTS13 protein in said sample by ultrafiltration; and stabilizing said ADAMTS13 protein by diafiltration exchange into a buffer comprising calcium ions and zinc ions ...

Purification method for divalent cation binding proteins by anion exchange chromatography

The present invention relates to a two-step method for the purification of divalent cation binding proteins with high yield and high purity on anion exchange resin materials, to divalent cation binding proteins obtainable by said method, and to a kit comprising means for carrying out said method. 112.-. (canceled)13. A method for the purification of a divalent cation binding protein comprising the steps of:(a) loading a first anion exchange resin material with the divalent cation binding protein in a loading buffer in the absence of divalent cations or at a low concentration thereof;(b) washing the loaded anion exchange resin material with a washing buffer comprising between 175 mM NaCl and 190 mM NaCl in the absence of divalent cations;(c) eluting the divalent cation binding protein with an eluant comprising a counter-anion to form an eluate containing the divalent cation binding protein;(d) supplementing the eluate obtained in step (c) with at least one divalent cation and increasing the pH;(e) loading a second anion exchange resin material with the supplemented eluate obtained in step (d); and(f) collecting the flow-through containing the divalent cation binding protein.14. The method according to claim 13 , wherein in step (d) the pH of the supplemented eluate is increased by at least 0.5 pH units.15. The method according to claim 14 , wherein the eluant in step (c) has a conductivity that is higher than the conductivity of the loading buffer in step (a) and higher than the conductivity of the washing buffer in step (b) and wherein the supplemented eluate in step (d) has a conductivity that is lower than the conductivity of the eluant in step (c).16. The method according to claim 13 , wherein the at least one divalent cation in step (d) is selected from the group consisting of Ca claim 13 , Be claim 13 , Ba claim 13 , Mg claim 13 , Mn claim 13 , Sr claim 13 , Zn claim 13 , Co claim 13 , Ni claim 13 , and Cu claim 13 , or combinations thereof.17. The method ...

PROTEIN PURIFICATION BY ANION EXCHANGE CHROMATOGRAPHY

The present invention relates to a two-step method for the purification of divalent cation binding proteins with high yield and high purity on anion exchange resin materials, to divalent cation binding proteins obtainable by said method, and to a kit comprising means for carrying out said method. 1. A method for the purification of a divalent cation binding protein comprising the steps of:(a) loading a first anion exchange resin material with the divalent cation binding protein in a loading buffer in the absence or low concentration of divalent cations, optionally followed by one to three wash steps;(b) eluting the divalent cation binding protein with an eluant comprising a divalent cation and a counter-anion to form an eluate containing the divalent cation binding protein;(c) diluting the obtained eluate pool, ((1) to optionally lower the conductivity), and increasing the concentration of the divalent cation;(d) loading a second anion exchange resin material with the eluate as obtained after step (c); and(e) collecting the flow-through containing the divalent cation binding protein.2. The method according to claim 1 , wherein after the loading step one or more washing steps (1) claim 1 , (2) and/or (3) with a washing buffer (1) claim 1 , (2) and/or (3) in the absence of a divalent cation but in the presence of a counter anion is/are performed.3. The method according to or claim 1 , wherein at least one of the loading buffer and/or washing buffer claim 1 , has/have a pH that is at least 0.5 pH units lower than the pH of the eluant of step (b) claim 1 , preferably wherein either the loading buffer and/or the buffer in wash step (2) has/have a pH that is at least 0.5 pH units lower than the pH of the eluant of step (b).4. The method according to any one or more of to claim 1 , wherein the eluant in step (b) has a conductivity that is higher than the conductivity of the loading buffer in step (a) and the optional washing buffer and wherein the supplemented. i.e. ...

PROTEIN PURIFICATION BY ANION EXCHANGE CHROMATOGRAPHY

The present invention relates to a two-step method for the purification of divalent cation binding proteins with high yield and high purity on anion exchange resin materials, to divalent cation binding proteins obtainable by said method, and to a kit comprising means for carrying out said method. 1. A method for the purification of a divalent cation binding protein comprising the steps of:(a) loading a first anion exchange resin material with the divalent cation binding protein in a loading buffer in the absence of divalent cations or at a low concentration thereof, and optionally washing the loaded anion exchange resin material with a washing buffer in the absence of divalent cations;(b) eluting the divalent cation binding protein with an eluant comprising a counter-anion to form an eluate containing the divalent cation binding protein;(c) supplementing the eluate obtained in step (b) with at least one divalent cation and increasing the pH;(d) loading a second anion exchange resin material with the supplemented eluate obtained in step (c); and(e) collecting the flow-through containing the divalent cation binding protein.2. The method according to claim 1 , wherein in step (c) the pH of the supplemented eluate is increased by at least 0.5 pH units.3. The method according to or claim 1 , wherein the eluant in step (b) has a conductivity that is higher than the conductivity of the loading buffer in step (a) and the washing buffer in step (a) claim 1 , in case a washing step is carried out claim 1 , and wherein the supplemented eluate in step (c) has a conductivity that is lower than the conductivity of the eluant in step (b).4. The method according to any of to claim 1 , wherein the at least one divalent cation in step (c) is selected from the group consisting of Ca claim 1 , Be claim 1 , Ba claim 1 , Mg claim 1 , Mn claim 1 , Sr claim 1 , Zn claim 1 , Co claim 1 , Ni claim 1 , and Cu claim 1 , or combinations thereof.5. The method according to anyone of to claim 1 , ...

VIRUS FILTRATION OF CELL CULTURE MEDIA

The invention relates to a method for removing a viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. The method comprises subjecting said preparation to filtration for at least about 24 hours through a virus filter having an effective pore size of maximum about 75 nm. Further, the invention relates to the use of a virus filter in filtration of at least about 24 hours, wherein the virus filter has an effective pore size of maximum about 75 nm for the removal of viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. In some embodiments the filtration according to the invention operates at a volumetric capacity of at least about 2000 L/m. Further, the invention relates to the use of a preparation, being a cell culture medium or at least a component of a cell culture medium obtainable according to method of the invention for cell culture; pharmaceutical, diagnostic and/or cosmetic preparations as well as in food preparations. 114.-. (canceled)15. An apparatus comprising a bioreactor and a virus filter , wherein the virus filter is inline of the medium feed line of the bioreactor , wherein the virus filter has an effective pore size of maximum 75 nm , and wherein the virus filter is for removing a viral contaminant from a preparation , said preparation being a cell culture medium or at least one component of a cell culture medium.16. The apparatus of claim 15 , wherein the bioreactor is a chemostat reactor claim 15 , a perfusion reactor claim 15 , or a fed batch reactor.17. The apparatus of claim 15 , wherein the filter comprises two or more filters arranged in series claim 15 , in parallel claim 15 , or both.18. The apparatus of claim 17 , wherein the filter comprises two filters arranged in parallel in a system of tubes comprising a Y-shaped junction and wherein each filter is in fluid communication with a branch of the Y-shaped junction and a ...

VIRUS FILTRATION OF CELL CULTURE MEDIA

The invention relates to a method for removing a viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. The method comprises subjecting said preparation to filtration for at least about 24 hours through a virus filter having an effective pore size of maximum about 75 nm. Further, the invention relates to the use of a virus filter in filtration of at least about 24 hours, wherein the virus filter has an effective pore size of maximum about 75 nm for the removal of viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. In some embodiments the filtration according to the invention operates at a volumetric capacity of at least about 2000 L/m. Further, the invention relates to the use of a preparation, being a cell culture medium or at least a component of a cell culture medium obtainable according to method of the invention for cell culture; pharmaceutical, diagnostic and/or cosmetic preparations as well as in food preparations. 1. A method for removing a viral contaminant from a liquid preparation comprising the steps of:supplying a liquid preparation to at least one replaceable virus filter having an effective pore size of 5-75 nm, wherein said filter is in fluid communication with a bioreactor; andremoving a viral contaminant by filtering the preparation through said virus filter, under pressure, for a minimum total run time of about 24 hours; andcollecting the filtered preparation;{'sup': '2', 'wherein the preparation comprises a cell culture medium or at least one component thereof; the virus filter is not replaced during said minimum run time; the pressure is from about 100 mbar to about 4000 mbar; and filtration operates at a volumetric capacity of at least about 2000 L/m.'}2. The method of claim 1 , wherein the bioreactor is a chemostat reactor claim 1 , a perfusion reactor claim 1 , or a fed batch reactor.3. The method of claim 1 , wherein ...

MEDIUM FOR THE PROTEIN-FREE AND SERUM-FREE CULTIVATION OF CELLS

A medium is described for the protein-free and serum-free cultivation of cells, especially mammalian cells, whereby the medium contains a proportion of soy hydrolysate. 115-. (canceled)16. A method for increasing productivity of a recombinant protein in a cell culture , the method comprising:culturing mammalian cells that express the recombinant protein in an animal protein-free and serum-free cell culture medium; andadding more than 10 wt. % ultrafiltered soy hydrolysate based on the total dry weight of the medium to the animal protein-free and serum-free cell culture medium, wherein at least 40% of the soy hydrolysate has a molecular weight of 500 daltons.17. The method of claim 16 , wherein at least 50% of the soy hydrolysate has a molecular weight of ≦500 daltons.18. The method of claim 16 , wherein at least 55% of the soy hydrolysate has a molecular weight of 500 daltons.19. The method of claim 16 , wherein the soy hydrolysate has an endotoxin content of <500 U/g.20. The method of claim 16 , wherein the mammalian cells are CHO cells or BHK cells.21. The method of claim 16 , wherein the recombinant protein is selected from the group consisting of: Factor II claim 16 , Factor V claim 16 , Factor VII claim 16 , Factor VIII claim 16 , Factor IX claim 16 , Factor X claim 16 , Factor XI claim 16 , Protein S claim 16 , Protein C claim 16 , activated forms of these factors claim 16 , and vWF.22. The method of claim 21 , wherein the recombinant protein is Factor VIII.23. The method of claim 16 , wherein the medium comprises an amino acid.24. The method of claim 23 , wherein the amino acid is selected from the group consisting of L-asparagine claim 23 , L-cysteine claim 23 , L-cystine claim 23 , L-proline claim 23 , L-tryptophan claim 23 , L-glutamine and mixtures thereof.25. The method of claim 16 , wherein the medium further comprises: 1 to 100 g/L synthetic minimal medium; 0.05-1 g/L glutamine; 0.0005-0.05 g/L ascorbic acid; 0.1-10 g/L NaHCO; 0.0005-0.05 g/L ...

OLIGOPEPTIDE-FREE CELL CULTURE MEDIA

The present invention relates to oligopeptide-free cell culture media comprising at least 0.5 mg/L of a polyamine and to methods for cultivating cells in said oligopeptide-free cell culture media comprising at least 0.5 mg/L of a polyamine. The invention also relates to methods for expressing at least one protein in a medium comprising at least 0.5 mg/L of a polyamine and to methods for producing at least one virus in a medium comprising at least 0.5 mg/L of a polyamine. 1. An oligopeptide-free cell culture medium , comprising at least 0.5 mg/L of a polyamine.2. The oligopeptide-free cell culture medium according to claim 1 , wherein the polyamine is selected from the group consisting of cadaverine claim 1 , putrescine claim 1 , spermidine claim 1 , spermine claim 1 , agmatine claim 1 , ornithine claim 1 , and combinations thereof.3. The oligopeptide-free cell culture medium according to claim 1 , wherein the polyamine is synthetically produced.4. The oligopeptide-free cell culture medium according to claim 1 , wherein the polyamine is present in the culture medium in a concentration ranging from about 0.5 to about 30 mg/L.5. The oligopeptide-free cell culture medium according to claim 1 , wherein the medium does not comprise oligopeptides having twenty or more amino acids.6. The oligopeptide-free cell culture medium according to claim 1 , wherein the medium does not comprise oligopeptides having three or more amino acids claim 1 , optionally comprising glutathione.7. The oligopeptide-free cell culture medium according to claim 1 , wherein the medium does not comprise oligopeptides having two or more amino acids claim 1 , optionally comprising glutathione and/or at least one stable form of glutamine.8. The oligopeptide-free cell culture medium according to claim 1 , wherein the medium is chemically defined.9. A method for cultivating cells claim 1 , comprising the steps of:{'claim-ref': {'@idref': 'CLM-00001', 'claim 1'}, '(a) providing an oligopeptide-free cell ...

FORMULATION OF SUGAR SOLUTIONS FOR CONTINUOUS ULTRACENTRIFUGATION FOR VIRUS PURIFICATION

The present invention provides a method for purification of a virus or virus antigen comprising providing a virus preparation and centrifugation of said virus preparation in a gradient of a sugar established by the addition of two or more buffered sugar layers of different concentration. The method leads to higher yields and reduces unwanted aggregation of the virus or virus antigen by increasing the volume of the peak pool. 134-. (canceled)35. A method for purification of a virus or a virus antigen , comprising (i) a first buffered sugar solution comprising a first physiological buffer and', '(ii) a second buffered sugar solution having a higher density than the first buffered sugar solution and comprising a second physiological buffer, which is the same or different from the first physiological buffer,', 'wherein the concentration of sugar in the first buffered sugar solution has a sucrose equivalent between 35% to 50% (w/w %) and the concentration of sugar in the second buffered sugar solution has a sucrose equivalent between 50% to 65% (w/w %);, 'providing a solution containing a sugar gradient by centrifuging in an ultracentrifuge at least'}adding a virus preparation to the sugar gradient and centrifuging in an ultracentrifuge comprising an ultracentrifuge rotor to obtain a peak pool, andextracting the peak pool to obtain the virus or viral antigen.36. The method of claim 35 , wherein the peak pool is extracted with a sucrose equivalent between 30% and 54% (w/w %) sucrose.37. The method of claim 35 , wherein at least one buffer has a concentration of between 5 mM to 50 mM.38. The method of claim 35 , wherein the step of centrifuging the virus preparation and buffered sugar solutions is performed with a relative centrifugation of at least 20 claim 35 ,000 g.39. The method of claim 35 , wherein the virus preparation is obtained by centrifugation without a preclarifier.40. The method of claim 35 , wherein said first buffered sugar solution comprises a sugar in a ...

Pegylation of recombinant blood coagulation factors in the presence of bound antibodies

Pegylation of recombinant blood coagulation factors in the presence of bound antibodies

The present invention relates to a proteinaceous construct comprising a blood coagulation factor, e.g., Factor VIII (FVIII), being bound to at least one water soluble polymer, including a poly(alkylene oxide) such as polyethylene glycol (PEG). Further the present invention relates to methods of preparing PEGylated blood coagulation factor, e.g., FVIII, in the presence of bound antibodies. The invention also relates to methods for prolonging the in vivo-half-life of blood coagulation factor, e.g., FVIII, in the blood of a mammal having a bleeding disorder associated with functional defects or deficiencies of blood coagulation factor, e.g., FVIII.

Pegylation of recombinant blood coagulation factors in the presence of bound antibodies

The present invention relates to a proteinaceous construct comprising a blood coagulation factor, e.g., Factor VIII (FVIII), being bound to at least one water soluble polymer, including a poly(alkylene oxide) such as polyethylene glycol (PEG). Further the present invention relates to methods of preparing PEGylated blood coagulation factor, e.g., FVIII, in the presence of bound antibodies. The invention also relates to methods for prolonging the in vivo-half-life of blood coagulation factor, e.g., FVIII, in the blood of a mammal having a bleeding disorder associated with functional defects or deficiencies of blood coagulation factor, e.g., FVIII.

Process for the separation of recombinant pro-factor IX from recombinant factor IX

Culture medium for protein-free and serum-free culture of cell

【課題】組換え細胞が、無血清もしくは無タンパク質様式で効率的に培養され得る、因子及び方法の提供。【解決手段】１０．３％と１５．６％との間の遊離アミノ酸含量および７．６％と１１．４％との間の総窒素含量を有するダイズ加水分解産物を含む、無動物タンパク質かつ無血清の細胞培養培地。【選択図】なし

Method of separating recombinant pro-factor IX from recombinant factor IX

Purification of vwf for increased removal of non-lipid enveloped viruses.

La presente invención proporciona métodos para purificar el factor de Von Willebrand (VWF) para la remoción incrementada de virus envueltos sin lípido.

Method for the purification of recombinant blood coagulation factor ix enriched in sulfated and/or phosphorylated molecules

Protein purification by anion exchange chromatography

The present invention relates to a two-step method for the purification of divalent cation binding proteins with high yield and high purity on anion exchange resin materials, to divalent cation binding proteins obtainable by said method, and to a kit comprising means for carrying out said method.

Methods of purifying recombinant ADAMTS13 and other proteins and compositions thereof

Provided herein are methods for purifying recombinant A Disintegrin-like and Metallopeptidase with Thrombospondin Type 1 Motif 13 (ADAMTS13) protein from a sample. The method comprises enriching for ADAMTS13 protein by chromatographically contacting the sample with hydroxyapatite under conditions that allow ADAMTS13 protein to appear in the eluate or supernatant from the hydroxylapatite. The methods may further comprise tandem chromatography with a mixed mode cation exchange/hydrophobic interaction resin that binds ADAMTS13 protein. Additional optional steps involve ultrafiltration/diafiltration, anion exchange chromatography, cation exchange chromatography, and viral inactivation. Also provided herein are methods for inactivating virus contaminants in protein samples, where the protein is immobilized on a support. Also provided herein are compositions of ADAMTS13 prepared according to said methods.

High molecular weight and low molecular weight fractions of von Willebrand factor

Sepn. of von Willebrand factor (vWF) into high and low mol. wt. fractions comprises binding vWF to an affinity column and carrying out elution at different salt concns. Also new are: (1) low mol. wt. fraction (A) of vWF, consisting mainly of dimers and tetramers and (2) high mol. wt. fraction (B) of at ≥ 50 (pref. ≥ 60)% increased activity (per mu g. of protein) in platelet aggregation compared with the physiological mixt. of both forms.

Purification of vwf for increased removal of non-lipid enveloped viruses

The present invention provides methods for purifying Von Willebrand factor (VWF) for increased removal of non-lipid enveloped viruses.

Method for the isolation and purification of vitamin K-dependent proteins

A method for the separation of vitamin K-dependent proteins from non-vitamin K-dependent accompanying proteins is described wherein the method is characterized in that at least anion exchange chromatography and optionally affinity chromatography is carried out as well. The method is suitable especially for the purification of Factor II, VII, IX, X as well as Protein S, Protein C and Protein Z. With the aid of the method according to the invention a vitamin K-dependent protein is obtained which is present at a purity of 95%.

PROCESS FOR PRODUCING PROTEINS

Purification of von willebrand factor by cation exchanger chromatography

The invention relates to a method for obtaining vWF, whereby vWf is bonded to a cation exchanger at a low salt concentration and vWf with high specific activity is obtained by fractional elution. The invention also relates to a preparation with purified vWf which can be obtained by this method.

Process for extracting high-purity von willebrand factor

The invention concerns a process for extracting high-purity von Willebrand factor wherein recombinant von Willebrand factor (rvWF) is chromatographically purified by anion exchange chromatography on a quaternary amino-type anion exchanger in a buffer solution comprising buffer substances and optionally salt. The buffer solutions are preferably free from stabilizers, amino acids and other additives. This process produces high-purity recombinant vWF which is free from blood plasma proteins, in particular factor VIII, and is physiologically active. The invention further concerns a pharmaceutical preparation which contains rvWF and is composed of multimers having high structural integrity.

Formulation of Sugar Solutions for Continuous Ultracentrifugation for Virus Purification

The present invention provides a method for purification of a virus or virus antigen comprising providing a virus preparation and centrifugation of said virus preparation in a gradient of a sugar established by the addition of two or more buffered sugar layers of different concentration. The method leads to higher yields and reduces unwanted aggregation of the virus or virus antigen by increasing the volume of the peak pool.

Method for purifying recombinant adamts13 and other proteins and compositions thereof

Medium for the protein-free and serum-free cultivation of cells

A medium is described for the protein-free and serum-free cultivation of cells, especially mammalian cells, whereby the medium contains a proportion of soy hydrolysate.

Substantially animal protein-free recombinat furin and methods for producing same

The present invention relates to recombinant furin (rFurin) and methods for producing rFurin. More specifically, the invention relates to substantially animal protein-free rFurin and methods for producing substantially animal protein-free rFurin.

Virus filtration of cell culture media

A method for removing a viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium is disclosed. The method comprises subjecting the preparation to filtration for at least about 24 hours through a virus filter having an effective pore size of maximum about 75 nm. The use of a virus filter in filtration of at least about 24 hours is disclosed where the virus filter has an effective pore size of maximum about 75 nm for the removal of viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium.

Procedure for recovery of the right vWF or factor VIII / vWF complex

Method for the purification of recombinant blood coagulation factor ix enriched in sulfated and/or phosphorylated molecules

High molecular and low molecular fractions of von willebrand factor

The invention provides high and low molecular weight fraction of von Willebrand Factor (vWF), which can be obtained by absorbing vWF to a heparin affinity support followed by eluting the vWF at differing salt concentrations.

Virus filtration of cell culture media

The invention relates to a method for removing a viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. The method comprises subjecting said preparation to filtration for at least about 24 hours through a virus filter having an effective pore size of maximum about 75 nm. Further, the invention relates to the use of a virus filter in filtration of at least about 24 hours, wherein the virus filter has an effective pore size of maximum about 75 nm for the removal of viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. In some embodiments the filtration according to the invention operates at a volumetric capacity of at least about 2000 L/m2. Further, the invention relates to the use of a preparation, being a cell culture medium or at least a component of a cell culture medium obtainable according to method of the invention for cell culture; pharmaceutical, diagnostic and/or cosmetic preparations as well as in food preparations.

Method of production of purified hepatitis a virus particles and vaccine preparation

The present invention provides methods of purification of Hepatitis A Virus from the supernatant of an infected cell culture and production of a preparation of purified HAV antigen. The present invention is also directed to an HAV vaccine composition comprising a preparation consisting of purified mature HAV particles in an amount sufficient to induce a protective immune response in a mammal.

Virus filtration of cell culture media.

La presente invención se refiere a un método para eliminar un contaminante viral de una preparación, siendo un medio de cultivo celular o al menos un componente de un medio de cultivo celular. El método comprende someter la preparación a una filtración durante al menos aproximadamente 24 horas a través de un filtro de virus que tiene un tamaño de poro efectivo de aproximadamente 75 nm máximo. Además, la presente invención se refiere al uso de un filtro de virus en una filtración de al menos aproximadamente 24 horas, en donde el filtro de virus tiene un tamaño de poro efectivo de aproximadamente 75 nm máximo para la eliminación de un contaminante viral de una preparación, siendo un medio de cultivo celular o al menos un componente de un medio de cultivo celular. En algunas modalidades la filtración de acuerdo con la presente invención opera en una capacidad volumétrica de al menos aproximadamente 2000 L/m2. Además, la presente invención se refiere al uso de una preparación, siendo un medio de cultivo celular o al menos un componente de un medio de cultivo celular que se puede obtener de acuerdo con el método de la presente invención para un cultivo celular; preparaciones farmacéuticas, de diagnóstico y/o cosméticas así como en preparaciones alimenticias.

Targeted angiogenesis

The invention relates to compositions, methods, and gene therapy reagents to promote or to inhibit angiogenesis in the treatment of peripheral vascular or cardiovascular diseases, utilizing a chimeric molecule comprising an angiogenic factor linked to a targeting molecule that specifically binds to a vascular endothelium.

Method for purifying recombinant ADAMTS13 and other proteins and compositions thereof

Provided herein are methods for purifying recombinant A Disintegrin-Iike and Metallopeptidase with Thrombospondin Type 1 Motif 13 (ADAMTS13) protein from a sample. The method comprises enriching for ADAMTS13 protein by chromatographically contacting the sample with hydroxyapatite under conditions that allow ADAMTS13 protein to appear in the eluate or supernatant from the hydroxylapatite. The methods may further comprise tandem chromatography with a mixed mode cation exchange/hydrophobic interaction resin that binds ADAMTS13 protein. Additional optional steps involve ultrafiltration/diafiltration, anion exchange chromatography, cation exchange chromatography, and viral inactivation. Also provided herein are methods for inactivating virus contaminants in protein samples, where the protein is immobilized on a support. Also provided herein are compositions of ADAMTS13 prepared according to said methods.

VIRUS FILTERING OF CELL CULTURE MEDIA

Process for extracting and purifying recombinant, non-lipidised osp-protein

A recombinant, non-lipidised OspC is disclosed, as well as a process for preparing Osp-proteins and vaccines which contain these recombinant proteins.

Method for purifying recombinant adamts13 and other proteins and compositions thereof

620988 Discloses a method for inactivating lipid-enveloped virus contaminants in a protein solution in the purification of a therapeutic protein composition, the method comprising: providing a protein solution comprising the therapeutic protein, immobilising the protein on a support, subsequently treating the protein (while it remains on the support) with a solvent-detergent mixture comprising a non-ionic detergent and an organic solvent, wherein the treatment results in the reduced formation of aggregates of the protein and lipid-enveloped virus contaminants are inactivated.

Isolation and purification method of proteins dependent from vitamin k

A process for the sepn. of vitamin=K dependent proteins from non-vitamin=K dependent proteins by the addn. of Ca<2+> to a protein-contg. soln., and contacting the soln. with an anion exchanger so that all proteins except the vitamin=K-dependent ones are adsorbed is claimed.

Virus filtration of cell culture media

Purification of von willebrand factor by cation exhcanger chromatography

The invention relates to a method for obtaining vWF, whereby vWf is bonded to a cation exchanger at a low salt concentration and vWf with high specific activity is obtained by fractional elution. The invention also relates to a preparation with purified vWf which can be obtained by this method.

Virus filtration of cell culture media

The invention relates to a method for removing a viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. The method comprises subjecting said preparation to filtration for at least about 24 hours through a virus filter having an effective pore size of maximum about 75 nm. Further, the invention relates to the use of a virus filter in filtration of at least about 24 hours, wherein the virus filter has an effective pore size of maximum about 75 nm for the removal of viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. In some embodiments the filtration according to the invention operates at a volumetric capacity of at least about 2000 L/m2. Further, the invention relates to the use of a preparation, being a cell culture medium or at least a component of a cell culture medium obtainable according to method of the invention for cell culture; pharmaceutical, diagnostic and/or cosmetic preparations as well as in food preparations.

Virus filtration of cell culture media

The invention relates to a method for removing a viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. The method comprises subjecting said preparation to filtration for at least about 24 hours through a virus filter having an effective pore size of maximum about 75 nm. Further, the invention relates to the use of a virus filter in filtration of at least about 24 hours, wherein the virus filter has an effective pore size of maximum about 75 nm for the removal of viral contaminant from a preparation, being a cell culture medium or at least a component of a cell culture medium. In some embodiments the filtration according to the invention operates at a volumetric capacity of at least about 2000 L/m 2 . Further, the invention relates to the use of a preparation, being a cell culture medium or at least a component of a cell culture medium obtainable according to method of the invention for cell culture; pharmaceutical, diagnostic and/or cosmetic preparations as well as in food preparations.

Complex suitable for carrying out a method of purifying pre-s hepatitis b surface antigen

ABSTRACT OF THE DISCLOSURE: There is disclosed a complex comprised of an insoluble polymer carrier to which monomeric human albumin is covalently bound and of a pre-S hepatitis B surface antigen bound in an elutable form to the monomeric human albumin by its pre-S(2)- and/or pre-S(1)-region. This complex may be used for therapeutic and diagnostic purposes and enables the rapid and efficient purification of pre-S-HBsAg by affinity chromatography.

Method for purifying recombinant adamts13 and other proteins and compositions thereof

Separation and purification process of proteins depending on k vitamin

Process for the isolation and purification of vitamin k dependent proteins

Purification method for divalent cation binding proteins on anion exchange resin

Disclosed is a method for the purification of a divalent cation binding protein comprising the steps of: a) loading an anion exchange resin material with the divalent cation binding protein in a loading buffer in the absence of divalent cations, and optionally washing the loaded anion exchange resin material with a washing buffer in the absence of divalent cations; and b) eluting the divalent cation binding protein with an eluant comprising at least one divalent cation to form an eluate containing the divalent cation binding protein; wherein the eluant in step (b) has a pH higher than the pH of the washing buffer in step (a), or, in case no washing step is carried out, of the loading buffer in step (a), wherein the pH of the eluant in step (b) is at least 0.5 pH units higher than the pH of the washing buffer in step (a), or, in case no washing step is carried out, of the loading buffer in step (a).

Factor VIII/vWF-complex and methods of purifying same

There is disclosed a method of recovering factor VIII/vWF-complex which is characterized in that factor VIII/vWF-complex from a protein solution is bound to a cation exchanger and is recovered by step-wise elution of factor VIII/vWF-complex, which particularly contains high-molecular vWF multimers, as well as a factor VIII/vWF-complex obtainable by means of cation exchange chromatography.

POLYDISPERSE NATIVE PSEUDOMONAS FLAGELLA (H) ANTIGENS (SUBJECTS) AND PROCEDURE FOR THEIR RECOVERY

Purification of von willebrand factor by cation exchanger chromatography

Method for producing mature VWF from VWF pro-peptide

The present invention relates to a method for producing a mature von Willebrand Factor (VWF) from von Willebrand Factor pro-peptide comprising the steps: immobilizing VWF pro-peptide on an ion exchange resin, incubating the immobilized VWF pro-peptide with furin to obtain immobilized mature VWF, and isolating mature VWF from the ion exchange resin by elution.

Virus filtration of cell culture media

Protein purification by anion exchange chromatography

The present invention relates to a two-step method for the purification of divalent cation binding proteins with high yield and high purity on anion exchange resin materials, to divalent cation binding proteins obtainable by said method, and to a kit comprising means for carrying out said method.

Medium for the protein-free and serum-free cultivation of cells

A medium is described for the protein-free and serum-free cultivation of cells, especially mammalian cells, whereby the medium contains a proportion of soy hydrolysate.

Process for the purification of recombinant alpha 1-antitrypsin involving a step of anion exchange chromatography

The present invention relates to a method for obtaining highly purified recombinant alphal -antitrypsin (rAAT) using anion exchange chromatography, wherein a composition comprising rAAT and at least one impurity originating from cultivation of cells used for generating of rAAT is loaded onto a column containing anion exchange material and the anion exchange chromatography is carried out with buffers containing phosphate ions and N-acetylcysteine.

method to produce mature von willebrand factor from von willebrand factor propeptide

método para produzir fator de von willebrand maduro a partir de pró-pertídeo de fator de von willebrand, e, uso de um fator de von willebrand maduro. a presente invenção diz respeito a um método para produzir um fator von willebrand compreendendo as etapas: - imobilizar pró-peptídeo de vwf em uma resina de troca de íon, incubando o pró-peptídeo de vwf imobilizado com furina para obter vwf maduro imobilizado, e -isolar vwf maduro da resina de troca de íon por eluição. method to produce mature von willebrand factor from von willebrand factor propeptide, and use of a mature von willebrand factor. the present invention relates to a method for producing a von willebrand factor comprising the steps: - immobilizing vwf propeptide on an ion exchange resin, incubating the immobilized vwf propeptide with furin to obtain immobilized mature vwf, and -isolate mature vwf from ion exchange resin by elution.

Targeted angiogenesis

The invention relates to compositions, methods, and gene therapy reagents to promote or to inhibit angiogenesis in the treatment of peripheral vascular o r cardiovascular diseases, utilizing a chimeric molecule comprising an angiogenic factor linked to a targeting molecule that specifically binds to a vascular endothelium.

Substantially animal protein-free recombinat furin and methods for producing same

Formulation of sugar solutions for continuous ultracentrifugation for virus purification

VON WILLEBRAND FACTOR PURIFICATION THROUGH CATIONIC EXCHANGE CHROMATOGRAPHY.

Procedimiento para la obtención de vWF, caracterizado porque se une el vWF con una concentración de sal <= 250 mM a un cambiador de cationes y, mediante elución fraccionada con una concentración de sal > 300 mM, se obtiene un vWF con una alta actividad específica que comprende especialmente multímeros de vWF de alto peso molecular. Procedure for obtaining vWF, characterized in that the vWF is joined with a salt concentration <= 250 mM to a cation exchanger and, by fractional elution with a salt concentration> 300 mM, a vWF with a high specific activity is obtained which especially comprises high molecular weight vWF multimers.

METHOD TO REMOVE A VIRUS WITHOUT A LIPID ENVELOPE FROM A SOLUTION

MÉTODO PARA REMOVER UM VÍRUS ENVELOPADO COM NÃO LIPÍDIO DE UMA SOLUÇÃO. A presente invenção fornece métodos para purificar fator Von. Willebrand (VWF) para remoção elevada de vírus envelopados com não lipídio. METHOD FOR REMOVING A NON-LIPID ENVELOPED VIRUS FROM A SOLUTION. The present invention provides methods for purifying Von factor. Willebrand (VWF) for high removal of non-lipid enveloped viruses.

Recombinant furin substantially free of animal protein and methods for producing the same

Composición que comprende furina recombinante sustancialmente libre de proteína animal que carece de los dominios transmembrana y citoplasmático y que comprende una actividad específica de al menos 300 U/mg de proteína, comprendiendo la composición proteína de la célula huésped en una concentración de menos de aproximadamente 1,0 ng de proteína/U de actividad de furina o ADN de la célula huésped en una concentración de menos de aproximadamente 0,4 pg de ADN/U de actividad de furina, y que carece esencialmente de proteínas contaminantes del suero, en la que la actividad de furina recombinante se mide en un péptido sintético corto que contiene la secuencia de reconocimiento dibásica unida a un grupo amino- metil-cumarina (AMC) fluorescente, que se libera después de la escisión (BOC-RVRR-AMC), por lo que una unidad de actividad se define como la liberación de 1 pMol de AMC por minuto a 30ºC. Composition comprising recombinant furin substantially free from animal protein lacking the transmembrane and cytoplasmic domains and comprising a specific activity of at least 300 U/mg protein, the composition comprising host cell protein at a concentration of less than about 1 0.0 ng protein/U furin activity or host cell DNA at a concentration of less than about 0.4 pg DNA/U furin activity, and essentially free of contaminating serum proteins, wherein recombinant furin activity is measured on a short synthetic peptide containing the dibasic recognition sequence linked to a fluorescent amino-methylcoumarin (AMC) group, which is released after cleavage (BOC-RVRR-AMC), thus that one unit of activity is defined as the release of 1 pMol of AMC per minute at 30°C.

Method of purification of hydrophobic proteins

The present invention relates to a method for obtaining highly purified hydrophobic proteins from cells by extraction using a buffer containing a detergent and removal of said detergent by hydroxyapatite (HA) column chromatography.

Method for purifying recombinant adamts13 and other proteins and compositions thereof

Provided herein are methods for purifying recombinant A Disintegrin-Iike and Metallopeptidase with Thrombospondin Type 1 Motif 13 (ADAMTS13) protein from a sample. The method comprises enriching for ADAMTS13 protein by chromatographically contacting the sample with hydroxyapatite under conditions that allow ADAMTS13 protein to appear in the eluate or supernatant from the hydroxylapatite. The methods may further comprise tandem chromatography with a mixed mode N cation exchange/hydrophobic interaction resin that binds ADAMTS13 protein. Additional optional steps involve ultrafiltration/diafiltration, anion exchange chromatography, cation exchange chromatography, and viral inactivation. Also provided herein are methods for inactivating virus contaminants in protein samples, where the protein is immobilized on a support. Also provided herein are compositions of ADAMTS13 prepared according to said methods. WO 2011/012726 PCT/EP2010/061192 1/2 FIG. 1 101 Concentration and buffer exchange by UF/DF (10-20 fold) (cut-off: 30 kDA) 106 102 r__ _ Concentration and buffer exchange Capture: by UF/DF with Dialyzer Hardware Anion Exchange Chromatography (cut-off: 10 kDA) (Edwards on ANX-Sepharose-FF low sub Lifesciences) 105 105 _ _ _ _ _ _ _ _ _ _ _ _ S/D Virus inactivation Virus removal by 30 minutes at 12-16°C Nanofiltration 0.2 p filtration before and (Planova 20 N) Asahi Kasei after treatment 103 107 Polishing (1a): Polishing 2: Hydroxyapatite Type l (Biorad) (incl concentration and formulation) E (non-binding mode) Cation Exchange Chromatography ( on Poros 50 HS (Applied Biosystems) 104 108 HU Polishing (1b): Mixed Mode Chromatography purified A13 BDS on Capto MMC (GE Healthcare) stored frozen at < -60 0 C

Protein and serum-free cell culture medium

A medium is described for the protein-free and serum-free cultivation of cells, especially mammalian cells, whereby the medium contains a proportion of soy hydrolysate.

Process for extracting and purifying recombinant, non-lipidised osp-protein

Beschrieben wird ein rekombinantes OspC, welches nicht-lipidiert ist, ein Verfahren zur Herstellung von Osp-Proteinen, sowie Impfstoffe, welche diese rekombinanten Proteine umfassen.

Purification method for divalent cation binding proteins on anion exchange resin

The present invention relates to a method for the purification of divalent cation binding proteins with high purity on an anion exchange resin material, to divalent cation binding proteins obtainable by said method, and to a kit comprising means for carrying out said method.