Hi All,
I'm performing a cell synchronization experiment in HeLa cells using a double thymidine block.
I would like to perform downstream experiments with cells at different phases of the cell cycle that require re-plating the cells, and I'm trying to determine at which point to trypsinize my cells.
I've been trying to find information about whether trypsin alters the cell cycle, or if cells continue to go through the cell cycle as normal during this process, but I haven't had much luck.
Any information is much appreciated!
Dear Melanie!
Please You look at this article:
https://www.biorxiv.org/content/10.1101/679928v1.full
Hi Melanie,
You can find some more details below mentioned weblink.
https://www.sigmaaldrich.com/technical-documents/articles/biology/cell-dissociation-with-trypsin.html
It may be useful for you.
Good Luck.
Hello Melanie
During trypsinization the cells detach from the substratum and change their morphology becoming spherical and cluster together. They become arrested in the G0/G1 phase of the cell cycle. Serum plays a very important role in the cell cycle. During trypsinization when serum is completely removed the cells are starved of nutrients which are present in the serum, and they freeze at that very point in the cell cycle.
In case of HeLa cells the cell cycle time is approximately 20 hours with G1 phase comprising half the cell cycle time which is most variable followed by the S phase. G2 phase is about 2-3 hours while the M phase is about 1 hour.
Trypsinization can be carried out at various time points after seeding till the 11th hour to obtain cells in the G1 phase, at 15-16 hour to obtain cells in the S phase, at 19th hour to obtain G2 population and at the 20th hour to obtain M population. I suggest you trypsinize the cells not in the first cell cycle but rather the second cell cycle.
Thank you.
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