I am having issues with reviving C6/36 WAlbB (C6/36 infected with Wolbachia from the mosquito Aedes albopictus ). Firstly they seems to be growing very slowly, secondly they are dying once split. I do trypsinise for 2 min followed by 2/3 gentle taps to dislodge cells from the surface. However, cells die in the subculture. I am using 1:1 ratio of Hepes-RPMI : Schneider's to support their growth. Could you please provide your insights on it?
Typically C6/36 cells do not need to be trypsinised, which could be why the cells are dying. Instead, we use a sterile cell scraper to scrape the cells off gently and then resuspend in fresh media. Also, it is typical for Wolbachia-infected insect cells to grow slower relative to Wolbachia-cleared cells since the cells are thought to undergo cellular and metabolic stress in the presence of the endosymbiont.
I agree with Tamanash. The cells might died due to Trypsinization.
Hello Dr. Bhattacharya, If we need to detach the cells from a culture plate (e.g., 96 or 24 well plates) I cannot use the cell scraper. If trypsin is not good for cell viability, what would you recommend me to use? Thank you. Tamanash Bhattacharya
Diana Giron-Ceron
I'd try something like PBS + EDTA (1-10mM) and gentle rocking to see if the cells come off. I haven't tried that on C6s personally, but it might work.
In my experience, I've not had to use Trypsin for C6/36 cells. I've always just tapped/pipetted and they would come off the plate. The recommendations from ATCC also are similar. Trypsinization could definitely harm the cells.
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