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Method for preparing porcine parvovirus inactivated vaccines

A technology for parvoviruses and inactivated vaccines, applied in biochemical equipment and methods, antiviral agents, viruses/bacteriophages, etc., can solve the problems of high labor intensity, hidden pollution, large difference between vaccine batches, etc., and improve ST Effects of cell density, increased virus titer, and stable quality

Inactive Publication Date: 2011-01-19
扬州优邦生物药品有限公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the production process adopts the traditional spinner bottle culture ST cell technology, that is, the inactivated porcine parvovirus vaccine is produced by inoculating the spinner bottle ST cells that have formed a single layer with the seed virus or inserting the seed virus at the same time when the cells are passaged. Each spinner bottle is an independent cell unit, so the cell quality, virus yield and titer of each bottle are different, which directly leads to the disadvantages of large batch-to-batch variation of vaccines and high endotoxin caused by hidden contamination, and at the same time, the labor intensity is high

Method used

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  • Method for preparing porcine parvovirus inactivated vaccines
  • Method for preparing porcine parvovirus inactivated vaccines

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1 Screening of suitable cell lines

[0024] The porcine parvovirus BJ-2 strain was inoculated in ST or IBRS-2 subculture cells, in DMEM medium, the culture conditions were controlled at pH 7.0-7.3, temperature 35-37°C, subcultured continuously, so that the virus adapted to the cells, and determined Virus reproduction titer, to reach the required virus content requirement ≥ 10 6.0 TCID 50 / ml to screen out suitable cell lines.

[0025] A suitable cell line should have the characteristics of good shape, stable and consistent state, and high toxin production. The specific cell standards are:

[0026] Cell morphology: use DMEM medium containing 10% fetal bovine serum in 5% CO 2 Incubate at 37°C in an incubator for 6 hours to adhere to the wall, and grow into a single layer after 40 hours. Observed under a microscope, the cells were irregular in shape.

[0027] Exogenous virus inspection: Inspect according to the appendix of the current "Chinese Veterinary Phar...

Embodiment 2

[0035] Embodiment 2 virus identification

[0036] Make BJ-2 strain virus on the suitable cell line screened out in embodiment 1, add in the DMEM medium that has 5~10% serum concentration, control culture condition is pH value 7.0~7.3, temperature 35~37 ℃ , continue to subculture, and at the same time identify the specificity, immunogenicity, virulence and other virus characteristics of the virus, and conduct gene sequence analysis to ensure that the virus is different from the original isolate in the specificity, virulence, immunogenicity and other aspects during the culture process. No significant changes have occurred.

[0037] (1) Specific identification of virus

[0038] Physical and chemical properties testing: the results show that the isolate is insensitive to lipid solvents, acids, and alkalis, and has strong thermal stability. These characteristics are consistent with the general characteristics of PPV.

[0039] Fluorescence inhibition test: Porcine parvovirus (PPV...

Embodiment 3

[0055] Example 3 Cell Line Culture

[0056] Use a square bottle to culture the selected ST cells or IBRS-2 cells recovered from liquid nitrogen. The culture conditions include: pH value 7.0-7.3, temperature 35-37°C, DMEM medium; culture for 48-72 hours, culture until the cells The concentration is 3~6×10 5 cells / ml, you can;

[0057] The cell line was cultured with microcarriers in a bioreactor, and the culture conditions were as follows: the medium was DMEM, and the cell seeding density was 1-2×10 5 cells / ml, the microcarrier concentration is 3-5g / L, the dissolved oxygen is 30%-60%, the pH value is 7.0-7.3, the temperature is 35-37°C, the rotation speed is 40-60r / min, and the cell concentration is 3-5 ×10 6 cells / ml. Wherein, the microcarrier is a spherical carrier Cytodex1 or a sheet carrier.

[0058] According to the growth characteristics of the cells and the size of the bioreactor volume, add fresh DMEM medium at a flow rate of 2 to 3 bioreactor volumes / day, and pump...

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Abstract

The invention discloses a method for preparing porcine parvovirus inactivated vaccines. The preparation method comprises the following steps of: recovery culture of ST cells or IBRS-2 cells, bioreactor micro-carrier culture of the ST cells or the IBRS-2 cells, continuous culture of bioreactor micro-carriers of the ST cells or the IBRS-2 cells, inoculated porcine parvovirus culture, collection of virus liquid, and virus inactivation for preparing the porcine parvovirus inactivated vaccines. The method for preparing the porcine parvovirus inactivated vaccines is a porcine parvovirus process for culturing the ST cells by adopting the bioreactor micro-carriers; the micro-carriers provide larger surface area, and can improve the density of the ST cells and the virus titer; the reactors can automatically monitor the growth of the cells or the optimal biochemical conditions during propagation of viruses; and the viruses produced on each batch of reactors have uniform titer, small batch and stable quality.

Description

technical field [0001] The invention relates to a preparation method of a vaccine, in particular to a preparation method of an inactivated porcine parvovirus vaccine. Background technique [0002] The production of porcine parvovirus inactivated vaccine using porcine testis (ST) passage cell line is a commonly used production process of porcine parvovirus inactivated vaccine. At present, the production process adopts the traditional spinner bottle culture ST cell technology, that is, the inactivated porcine parvovirus vaccine is produced by inoculating the spinner bottle ST cells that have formed a single layer with the seed virus or inserting the seed virus at the same time when the cells are passaged. Each spinner bottle is an independent cell unit, so the cell quality, virus yield and titer of each bottle are different, which directly leads to the disadvantages of large batch-to-batch variation of vaccines and high endotoxin caused by hidden contamination, and at the same...

Claims

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Application Information

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IPC IPC(8): A61K39/23C12N7/00A61P31/20
Inventor 李玉和史春云刁文华潘杰傅元华
Owner 扬州优邦生物药品有限公司
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