Method for preparing novel influenza vaccine
A technology for influenza and vaccines, which is applied in the field of new inactivated influenza vaccines, and can solve problems such as the source of influenza vaccine safety culture medium
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Embodiment 1
[0011] 1) Influenza virus subtypes A1, A3 and B were respectively diluted with virus culture medium at a ratio of 1:10 -4 Separately inoculate Vero cells that have grown to a single layer, and culture them at 35°C until the cytopathic pathology (CPE) reaches +++~++++++, then harvest the virus liquid containing influenza virus; the virus culture liquid The ingredients are:
[0012] MEM medium;
[0013] 1% bovine serum albumin; 5μg / ml trypsin;
[0014] Use HEPES to adjust the pH value of the solution to 7.0;
[0015] 200 μg / ml neomycin.
[0016] 2) The influenza virus in the virus liquid harvested in 1) was inactivated with 1:1000 formaldehyde solution, and the inactivated virus antigens were purified separately by chromatography, and purified by DEAE Streamlin chromatography, and the chromatography conditions were: : DEAE Streamline basic buffer: pH6.8, 0.01Mol / L PBS buffer, 0.01Mol / L sodium chloride for elution; obtain virus stock solution;
[0017] 3) Detect the antigen ...
Embodiment 2
[0020] 1) Influenza virus subtypes A1, A3 and B were respectively diluted with virus culture medium at a ratio of 1:10 -7 Separately inoculate Vero cells that have grown to a monolayer, and culture them at 33°C until the cytopathic pathology (CPE) reaches +++~++++++, and harvest the virus liquid containing influenza virus respectively; the virus culture liquid The ingredients are:
[0021] MEM medium;
[0022] 0.01% bovine serum albumin; 20μg / ml trypsin;
[0023] Use 5% sodium bicarbonate solution to adjust the pH value of the solution to 7.5;
[0024] 20 μg / ml neomycin.
[0025] 2) Inactivate the influenza virus in the virus liquid harvested in 1) with a 1:1000 β-propiolactone solution, and purify the inactivated virus antigens separately by chromatography, using DEAE Sepharose FF chromatography Purification, chromatographic conditions: Basic buffer of DEAE SepharoseFF: pH8.0, 0.2Mol / L Tris buffer, elution with sodium chloride concentration of 1Mol / L; obtain virus stock s...
Embodiment 3
[0029] 1) Influenza virus subtypes A1, A3 and B were respectively diluted with virus culture medium at a ratio of 1:10 -1 Separately inoculate Vero cells that have grown to a monolayer, and culture them at 34°C until the cytopathic pathology (CPE) reaches +++~++++++, and harvest the virus liquid containing influenza virus respectively; the virus culture liquid The ingredients are:
[0030] MEM medium;
[0031] 0.1% bovine serum albumin; 2μg / ml trypsin;
[0032] Use 6% sodium bicarbonate solution to adjust the pH value of the solution to 8.0;
[0033] 500 μg / ml neomycin.
[0034] 2) inactivate the influenza virus in the virus liquid harvested in 1) with a 1:10000 formaldehyde solution, and purify the inactivated virus antigens separately by chromatography, and purify by Sepharose 4FF chromatography, and then chromatographically Conditions: Sepharose 4FF elution buffer is pH7.5, 0.1Mol / L PBS buffer, containing 0.1Mol / L sodium chloride; obtain virus stock solution;
[0035] 3)...
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