Optimization of Lentivirus Production Process for 293T Cells
With the rapid development of cell therapy,large-scale lentiviral production has become a bottleneck in the process chain;thus,optimizing the production process of CAR lentiviral vectors with high titer and high purity of 293T becomes crucial.This study aimed to optimize the packaging of lentiviral 293T adherent cells to achieve time-saving,cost-saving,and improved lentiviral packaging.At the same time,the optimization of lentiviral vectors was carried out to explore the factors affecting the growth of suspension cell clusters.The 293T adherent cells were domesticated into suspension culture by fast and slow domestication.Their cell morphology,cell density,cell viability,lentiviral packaging ability,and stable consistency after cryopreservation and resuscitation were compared to screen out the optimal suspension domestication conditions.Comparative cell clumping growth was studied by adjusting Ca2+concentration and EDTA addition.The findings showed that the serum-free medium OPM-293 CD05 Medium could be quickly domesticated into 293T suspension cells from 293T adherent cells,and that these suspension cells could be produced with lentiviral titers that were better than the adherent cells'packaging titers(*P＜0.05).Ca2+concentration affects the size of cell clusters.The addition of EDTA effectively separates and disperses unnecessary cell clusters.In summary,the experiment's findings demonstrated that serum-free OPM-293 CD05 Medium could quickly domesticate conventional 293T adherent cells into suspension cells.Within a certain range,the higher the concentration of Ca2+,the larger the agglomerates and particle size,and the higher the addition of EDTA,the smaller the agglomerates and particle size.This provides a theoretical framework for the optimization of suspension culture conditions and the lentiviral vector packaging process.It also establishes a theoretical framework for the scale-up and manufacturing of in vitro cell culture,which has some practical applications.
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