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Method for separating and purifying blood coagulation factor VIII

A coagulation factor, separation and purification technology, applied in the field of separation and purification of coagulation factor VIII, can solve problems such as poor specificity

Inactive Publication Date: 2014-06-18
新疆德源生物工程有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Heparin is less expensive, but has poor affinity and specificity

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0009] Preparation of immunoaffinity chromatography column: Wash 2g of CNBr-activated sepharose 4B Fast Flow with 400ml of 10mmol / L HCl, then slowly add to the chromatography column, keep the outflow port open, and the gel will settle naturally; at this time, slowly add 2 %CaCO 3 , circulated 3 times, after the solution was drained, the anti-coagulation factor VIII antibody was filtered through a 0.22um bacterial filter and added to the column, and then 0.1mol / L NaHCO 3 Sufficient dialysis; after sufficient coupling, wash to effluent OD 280 0, and then fully wash the column with 200ml 0.01mol / L Tris-HCl, pH7.0 buffer to seal the uncoupled active site on the gel; use 3 times the volume of 0.01mol / L Tris-HCl, pH7. 0,3mmol / LCaCl 2 and 3 cycles of 0.05 mol / L Tris-HCl, pH 7.0, and 6 mmol / L EDTA to wash the chromatography column for 5 cycles.

[0010] Effect of Different Reaction Conditions on Separation Effect

[0011] Final effect of the present invention is compared with trad...

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Abstract

The invention discloses a method for separating and purifying a blood coagulation factor VIII. The method mainly comprises the steps of adsorbing a cryoprecipitate through DEAE (Diethyl-Aminoethanol)-Spherodex; then, carrying out immunoaffinity chromatographic purification; removing a blood coagulation factor VIII antibody by using DEAE agarose gel. The method is characterized by comprising the steps of firstly adsorbing 2% of CaCO3 on an affinity column, and then, adsorbing the blood coagulation factor VIII antibody on the affinity column; during affinity chromatography, uniformly stirring the blood coagulation factor VIII in a buffer solution, and eluting with an elution buffer solution to obtain a purified blood coagulation factor VIII. By using the method, the production cost can be reduced, the direct extraction of the blood coagulation factor VIII in plasma becomes possible, and the traditional separation method with low yield and low purity is prevented from being used.

Description

technical field [0001] The invention relates to a method for separating and purifying blood coagulation factor VIII, which belongs to the field of biological separation materials. Background technique [0002] Bioseparation ligands are a class of materials designed for the isolation and purification of biologically relevant products. This process is widely used in the separation of pharmaceutical industry, fermentation industry, biological products, blood products, vaccines and other products. When purifying products, the advantages of good separation effect and high product purity can be achieved by using the bioseparation ligand process. Especially when the content of the target product that needs to be separated and purified from plasma is low, this separation technology is particularly important. [0003] Blood coagulation factor Ⅷ is made from healthy human plasma, which is separated, purified, virus-removed and inactivated, and freeze-dried. The preparation can corr...

Claims

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Application Information

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IPC IPC(8): C07K14/755C07K1/22
CPCC07K14/755
Inventor 吕献忠苏峰
Owner 新疆德源生物工程有限公司