I'm working with a 293T tissue culture. I've been trying to seed 24-well plates, but I've been having trouble keeping the cells adhered to the bottom. I've worked with them before and never had issues like this, but when I try changing the media, the cells come off from the bottom of the well. I thought it was because it wasn't confluent enough, but I tried changing the media today of an almost over confluent plate, and I still had some issues. I've tried adding the media as slowly as possible. Can anyone think of any solutions?
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elpollodiablo
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Posted 13 October 2009 - 11:49 PM
melody89, on Oct 13 2009, 10:28 PM, said:
I'm working with a 293T tissue culture. I've been trying to seed 24-well plates, but I've been having trouble keeping the cells adhered to the bottom. I've worked with them before and never had issues like this, but when I try changing the media, the cells come off from the bottom of the well. I thought it was because it wasn't confluent enough, but I tried changing the media today of an almost over confluent plate, and I still had some issues. I've tried adding the media as slowly as possible. Can anyone think of any solutions?
Preheat the media, trypsinize them for a very short period of time (you might even want to use 0.0025% trypsin instead of 0.05%). When you have them in the incubator, dont put the lid completely on to allow better ventilation. I've had plenty of issues with these cells when using them in well-plates, whereas they grow great in flasks. But these tricks seem to help. Good luck!
