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
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[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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