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Method for producing rabies vaccine from cef cells suitable for sheet carrier bags

A technology of rabies vaccine and sheet-like carrier, which is applied in the field of CEF cell production of rabies vaccine and cell culture, can solve the problem of difficulty in beating with trypsin, and can solve the problem of DNA residue in host cells, good nutrient and gas exchange, and convenient operation. Effect

Active Publication Date: 2020-12-25
WUHAN CEKG TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] In addition, when the existing microcarriers are used for cell culture, tidal movement is often used for mass transfer and oxygen transfer. It is difficult to add pancreatin to internal digestion; the tidal reactor of Yisco company transfers mass and oxygen through ebb and flow. It pumps the liquid out through a peristaltic pump, and then another peristaltic pump pumps the liquid in. in; it is a static exchange, less efficient than dynamic

Method used

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  • Method for producing rabies vaccine from cef cells suitable for sheet carrier bags
  • Method for producing rabies vaccine from cef cells suitable for sheet carrier bags

Examples

Experimental program
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Effect test

preparation example Construction

[0036] (2) Preparation of CEF cells

[0037] Take 9-10 day-old SPF chicken embryos to prepare chicken embryo fibroblasts, namely CEF cells;

[0038] (2) Primary culture of CEF cells in sheet carrier bags

[0039] (2.1) Aseptically pump the sterile-filtered DMEM medium containing 10% newborn bovine serum into the sheet carrier bag, and place the sheet carrier bag in a swing bed to balance overnight, wherein, the sheet carrier bag culture conditions The temperature is 37°C, pH 7.2, dissolved oxygen 50%; the swing bed angle of the swing bed is 6-9°, and the speed is 10-30RPM;

[0040] (2.2) Insert the CEF cells prepared in (1) into the sheet carrier bag in (2.1), the inoculation density is 4-8×E5 / ml, and the swing bed angle is adjusted to 2-3°, the speed Adjust to 5RPM, and after 15min-2h, the cells are gradually attached to the sheet carrier; after 8-10h, adjust the swinging bed angle to 6-9°, and the speed is 10-30RPM;

[0041] (3) Access to rabies vaccine virus

[0042] Th...

specific Embodiment

[0058] (1) The purpose of the experiment: to verify the cultivation of CEF cells in the sheet carrier bag; the sheet carrier bag can be digested and amplified for transfer; the aG strain rabies vaccine virus that has been passed on the CEF chicken embryo cells for 2-3 generations can propagate the virus.

[0059] (2) Experimental materials:

[0060] One 3L carrier culture bag. 2 50L fluid storage bags. 1 temperature control swing bed. 1 set of SKC300 control system. 4 50ml beakers. 2 5L beakers. 1 glass rod. 1 capsule filter. 10 500ml blue cap bottles. 1 peristaltic pump. 5ml pipette. 10ml pipette. Electric pipette gun. square bottle. microscope. 1ml ep tube. cultured in a 5% CO2 incubator. Ultra clean bench. centrifuge. Hemocytometer. Cooling box etc.

[0061] 2 bottles of 0.25% trypsin, sodium bicarbonate, 1 bucket of DMEM medium, purified water, Nacl, Kcl, KH2PO4, Na2HPO4, 2 bottles of calf serum (CCS), 15ml centrifuge tube, thermometer, alcohol, trypan b...

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Abstract

The invention discloses a method for producing rabies vaccine by CEF cells suitable for sheet carrier bags, which comprises the following steps of: (1) preparing CEF cells; (2) primary culturing CEF cells in sheet carrier bags; (3) accessing rabies vaccine virus; (4) harvesting rabies vaccine virus solution to obtain rabies vaccine stock solution. According to the method, primary CEF cells cultureis directly carried out by adopting sheet carrier bags, and then rabies virus is inoculated, so that primary CEF cells required by large-scale culture can be cultured, and at the beginning of the production of rabies vaccine virus, namely SPF chicken embryos can be purchased directly and processed for current use at the beginning of cell culture without creating a database, the operation is convenient, time and labor are saved; and the problem of DNA residue of host cells is solved by adopting primary CEF cells to produce rabies virus vaccine. In addition, sheet carrier bags are used for carrying out mass transfer and oxygen transfer by swinging back and forth or left and right, and the static and dynamic combination of the swinging bed in the culture process is combined, so that the masstransfer and oxygen transfer efficiency is high.

Description

technical field [0001] The invention relates to the field of life sciences, especially cell culture and virus propagation. Specifically, it relates to a method for producing rabies vaccine from CEF cells suitable for sheet carrier bags. Background technique [0002] At present, most of the equipment used in the production of rabies vaccines and other products by CEF cells in the market is produced by glass or stainless steel bioreactors. However, when using glass and stainless steel bioreactors for production, a large number of spinner bottle passaging and digestion work and a large space are required in the early stage of cell proliferation and seed cell preparation. The production of the sheet carrier bag is to use a control system to control a swing bed reactor, place the sheet carrier bag on the swing bed, connect the gas control pipeline, and connect the cells with a square bottle, so that the cells can be placed in the swing bed. High-density proliferation and inocul...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00A61K39/205A61P31/14C12R1/93
Inventor 王树华甄宝贵李驰吴菲菲
Owner WUHAN CEKG TECH CO LTD
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