Serum-free suspension domestication method of Vero cells

Abstract

The invention relates to the field of biology. A serum-free suspension domestication method of Vero cells comprises the steps of step I, performing adherent culture on the Vero cells until the convergence rate reaches 80%, discarding supernatant, adding D-PBS free from calcium and magnesium ions, and performing washing; step II, discarding the D-PBS, adding pancreas enzymes, discarding trypsinization liquid, remaining a few pancreas enzymes which are enough to cover cells, performing incubation until the cells become round or performing incubation for definite time, and patting the bottom of aflask to dissociate cells; step III, performing re-suspension with a Vero-S suspension serum-free culture medium; step VI, discarding the supernatant, and performing re-suspension with Vero-S suspension serum-free culture medium; step V, diluting the density of the cells, performing inoculation to a disposable shake flask, and performing culture; step VI, when the density of the cells reaches 3*10<6> cells/ml, performing cell passage; and step VII, once the density of the cells can reach 3*10<6> cells/ml or above in 48h for continuous 10 generations or above and the motility rate of the cellsis not less than 95%, thinking that the cells are adapted to suspension culture.

Description

technical field [0001] The invention relates to the field of biology, in particular to cell culture. Background technique [0002] At present, for the cultivation of Vero cells, adherent culture is mostly used, but due to the limited attachment area that the culture system can provide, the number of proliferation and amplification is limited, and the culture process is difficult to scale up. Contents of the invention [0003] The object of the present invention is to provide a serum-free suspension acclimatization method of Vero cells, thereby solving the above-mentioned technical problems. [0004] The serum-free suspension domestication method of Vero cells is characterized in that: [0005] Step 1: Adhesively culture Vero cells until the confluency reaches 80%, discard the supernatant, and add D-PBS without calcium and magnesium ions to wash; [0006] Step 2: Discard D-PBS, add trypsin, shake at room temperature for a while, discard trypsin digestion solution, leave a...

AI Technical Summary

Problems solved by technology

[0002] At present, for the cultivation of Vero cells, adherent culture is mostly used, but due to the limited attachment area that the culture system can provide, the number of proliferation and expansion is limited, and the culture process is difficult to scale up.