Research on Serum-free Suspension Domestication of L929 Cells and its Application in Chlamydia abortus Inactivated Vaccine
In order to provide a suspension cell culture method for large-scale production of Chlamydia abortus,this study decreasing serum concentrations,adherent L929 cells adapted to suspension culture gradually,and became completed suspended,finally we get a L929 cell lines can cultured in serum-free suspension.Chlamydia abortus was inoculated on L929 suspension cells,and the best culture method was selected by adding different infection-promoting agents to increase the amount of Chlamydia abortus.Chlamydia abortus inactivated vaccine was prepared,and Balb/c mice were immunized with the vaccine at doses of 25 μL,50 μL and 100 μL,and the immunoprotective effect was detected after challenge.The results showed that when the inoculation density of the suspension cells was 5×105cells/mL,after 72 hours of culture,the cell concentration reached about 7×106 cells/mL,and the cell viability was more than 95%.When the cell density was 6×106cells/mL,the infection amount of pathogens was 5.5×104 IFU/mL,the addition of lipofectamine 2000 was 1.7 μL/mL and the concentration of actinomycetone was 0.4 μg/mL,the amount of pathogens was amplified by 52 times.After immunized with the suspended cell Chlamydia abortus inactivated vaccine,the antibody level in the mice was increased with the increase of immunization dose,and the level of splenic lymphocyte proliferation and the content of IFN-γ and IL-2 in 50 μL and 100 μL vaccine groups were much higher than those in 25 μL vaccine group and PBS group.The immunized mice were challenged,the genome content of Chlamydia abortus in spleen,duodenum and uterus of mice inoculated with vaccine was much lower than that of mice without vaccination.The uterus of mice injected with PBS and 25 μL of vaccine showed different degrees of necrosis and inflammation,while the uterus of the remaining two groups of mice had no abnormal changes.The results showed that a L929 suspension culture cell line was successfully acclimated in this experiment,and the inactivated Chlamydia abortus vaccine prepared from L929 suspension culture had strong immune effect.Inoculating 50 μL and 100 μL of the inactivated vaccine can have a protective effect on mice to a certain extent.The successful domestication of the L929 cell suspension culture strain provides a scientific basis for the large-scale preparation of vaccines.
adherent L929 cellsuspension culture and acclimationChlamydia abortusinactivated vaccineimmunological evaluation
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维普
