Method for extracting rabies virus

A rabies virus and molecular sieve technology, applied in the direction of microorganism-based methods, viruses, virus/bacteriophage, etc., can solve the problems of finished product preparation obstacles, restricted scale, and inability to industrialize production, achieve small fluctuations in antigen content, break production bottlenecks, The effect of rationalization of the production process

Inactive Publication Date: 2008-09-24
LIAONING YISHENG BIOLOGY PHARMACY
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] 1) There is a large difference in quality indicators between different batches, which has a great impact on the stability of biological products
[0004] 2) Removal of residual DNA becomes difficult
[0005] 3) The content of residual host protein is too high, and the side effects of clinical manifestations are relatively large
[0006] 4) After the original solution is prepared, the antigen content fluctuates in a wide range, and the preparation of the finished product in the later stage will cause certain obstacles
[0007] 5) It affects the product quality of biological products and restricts the scale of industrial production and extraction
However, the chromatography packing material is expensive and cannot be used in industrial production, and the loss of the antigen needs to be recovered, and the stability of the antigen needs to be investigated

Method used

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  • Method for extracting rabies virus
  • Method for extracting rabies virus
  • Method for extracting rabies virus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] 1 Use a hollow fiber ultrafiltration column with a molecular weight cut-off of 750KD or 500KD to concentrate the virus harvest liquid and remove some impurities

[0027] First, 10 L of the hollow fiber ultrafiltration column was equilibrated with 10 mmol / L PBS-0.15M NaCl buffer solution (pH 7.6). 40L virus harvest liquid was loaded and concentrated, the hollow fiber ultrafiltration column model was UFP-750-E6A, and the membrane area was 2800cm 2 , with a molecular weight cut-off of 750,000 Daltons. After the virus harvest solution was concentrated to 0.4L, 10 mL of the virus concentrate and filtrate were sampled respectively. Then use 10mmol / L PBS-0.15M NaCl buffer solution (pH7.6) to add and elute 8L of equilibrium liquid. Collect a sample of the filtered submembrane liquid every 0.4 L, and take 10 mL of the virus concentrated liquid sample at the same time. Detection of the protein concentration of the collected fractions showed that the removal rate of impurity pr...

Embodiment 2

[0063] 1. Utilize the ultrafiltration membrane with a molecular weight cut-off of 750KD or 500KD to concentrate the virus harvest liquid and remove some impurities

[0064] The operation process and parameters are the same as in Step 1 of Embodiment 1, so they are omitted.

[0065] 2. Extract the sample obtained in step 1 with molecular sieve gel 6% cross-linking degree high-flow agarose gel filtration (SepHarose 6FF)

[0066] Prepare one chromatographic column as mentioned in Example 1 or cut the chromatographic column into four in series to form quadruple columns, then each chromatographic column is 26×175mm, and the volume of each column is 32.5mL. It is best to use four columns connected in series to form a quadruple column.

[0067] The rest of the operation process is the same as Step 3 of Embodiment 1, so it is omitted.

[0068] 3. Extract the sample obtained in step 2 with anion exchange gel high flow rate tetravalent amino agarose (Q SepHarose FF)

[0069] The oper...

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Abstract

The invention provides a method for extracting rabies virus, and is to solve the defects that great discrepancy of quality indices of different batches occurs; removal of remaining DNA becomes difficult; the protein content of the remaining host is too high; and great side effects appear clinically when the single method of molecular sieve gel chromatography is adopted for extracting rabies virus vaccine. The essential of the invention is that a hollow fiber ultrafiltration column or ultrafiltration membrane with the molecular weight cut-off of 750KD or 500KD is used to condense and partially purify harvested liquid of virus; anion exchange chromatography or molecular sieve gel chromatography is adopted to separate and purify samples; molecular sieve gel chromatography or anion exchange chromatography is adopted to separate and purify samples got in step (2). The method for extracting rabies virus has the characteristics of great productive capacity, high product quality, excellent batch stability, being remarkably effective in removal of remaining DNA and HCP, and reducing the potential safety hazard of vaccine.

Description

technical field [0001] The invention relates to a method for extracting rabies virus using biotechnology. Background technique [0002] At present, the extraction of rabies virus vaccine generally adopts a single method of molecular sieve gel chromatography, which has the following defects: [0003] 1) There is a large difference in quality indicators between different batches, which has a great impact on the stability of biological products. [0004] 2) The removal of residual DNA becomes difficult. [0005] 3) The content of residual host protein is too high, and the side effects of clinical manifestations are relatively large. [0006] 4) After the stock solution is prepared, the antigen content fluctuates in a wide range, and the preparation of the finished product in the later stage will cause certain obstacles. [0007] 5) It affects the product quality of biological products and restricts the scale of industrial production and extraction. [0008] For this reason,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/02C12R1/93
CPCC12N2760/20151C12N7/00
Inventor 胡金东张译李伟刘伟李艳徐静
Owner LIAONING YISHENG BIOLOGY PHARMACY
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