Preparation of diploid cell attenuated live vaccine from measles long-47 strain and preparation process thereof
A diploid cell and attenuated live vaccine technology, applied in the field of vaccine preparation, can solve the problems of contamination allergic reaction, production efficiency and low quality, achieve less residual cells, improve vaccine harvest rate, vaccine quality, and harvest rate high effect
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Embodiment 1
[0018] 1) Passage human embryonic lung diploid cells (2BS) to the cell factory at a ratio of 1:4, the pH of the E-MEM culture medium is 7.2, the content of bovine serum is 10% (ml / ml), and the concentration is 3% The content of glutamine is 1%, and the amount added is 30% of the volume of the cell factory on each layer; at the same time, the multiplicity of infection of Chang-47 measles virus `1:500 is added to the cell factory, and the cells and viruses are cultivated at 33 °C;
[0019] 2) When the cells have 50-60% lesions, discard the liquid in the cell factory, wash the cell surface with 2-3 times the HanKs' solution or PBS solution of the culture medium, shake fully and empty the washing solution; add 199 vaccine solution, the amount added is 25-40% of the volume of the cell factory in each layer, continue to cultivate at 33°C, harvest the virus solution, and repeat three times;
[0020] 3) Store the virus liquid harvested three times at (2~8)°C. After passing the sterili...
Embodiment 2
[0024] 1) Passage human embryonic lung diploid cells (KMB-17) to the cell factory at a ratio of 1:3, the pH of the E-MEM culture medium is 7.4, the content of bovine serum is 12% (ml / ml), and the concentration is 3 % glutamine content is 1%, and the amount added is 35% of the volume of each cell factory; at the same time, Chang-47 strain of measles virus is added to the cell factory at a multiplicity of infection of 1:1000, and the cells and viruses are cultivated at 33°C;
[0025] 2) When the cells have 50-60% lesions, discard the liquid in the cell factory, wash the cell surface with 2-3 times the HanKs' solution or PBS solution of the culture medium, shake fully and empty the washing solution; add 199 vaccine solution, the amount added is 30% of the volume of the cell factory in each layer, and the culture is continued at 33°C; repeat three times to harvest the virus solution;
[0026] 3) Store the virus liquid harvested three times at (2~8)°C. After passing the sterility t...
Embodiment 3
[0030] 1) The human diploid cell (2BS) strain is subcultured at a ratio of 1:2 to the cell factory for use. The pH of the E-MEM growth medium is 7.6, the content of bovine serum is 14% (ml / ml), and cultured at 37°C 3 days, when the cells grow into a uniform and dense monolayer;
[0031] 2) Infect cells with Chang-47 strain of measles virus at a multiplicity of infection of 1:100. The pH of the maintenance medium E-MEM culture medium used is 7.4, the content of bovine serum is 2% (ml / ml), and the concentration is 3%. The aminoamide content is 1%, and the addition amount is 30% of the volume of each cell factory;
[0032] 3) When 50% of the cells are diseased, discard the liquid in the cell factory, wash the cell surface with HanKs' solution or PBS solution equivalent to 2~3 times, shake well, drain the washing solution, add 199 vaccine solution, Continue to culture at 33°C, and start to harvest the virus liquid when 75% of the cytopathic changes appear, and repeat three times;...
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