Help with clonogenic survival assay for PC3 prostate cancer cell line - HELP! (Jul/27/2008 )
Does any one have a good protocol and advice for clonogenic survival assays with PC3 Prostate Cancer Cell Line?
They strangely don't seem to stick very well on petri dishes even after treatment with drug at very low concentrations (within micromolar range) anyone has any idea why?
Please let me know! Thanks!
Hi they are weird cells aren't they. We find they need about 2 days to sit down on a TC dish. Sorry can't help with the assay.
yes they are really wierd cells- but when they stick in a flask, they seem to be alittle more difficult to remove even with 0.01% pronase/trypsin,
The problem was that for microscopy in chamber slides, they didn't seem to stick to the slide when the chambers were removed- but perhaps you're right, leave them for abit longer and they'll be stuck better.
Thanks for your advice! Much appreciated
For trypsinising, I leave them in trypsin in the incubator for a few minutes. They don't stick to glass very well.
Hmm yes that'd be an excellent idea! thanks! I've heard some reccomendations that perhaps when washing the cells, dulbecco's PBS -/- (no MgCl2/CaCl2) could possibly make the difference in preventing the cells from falling off- I realised I used 0.9% Saline solution- perhaps the pH buffering qualities of PBS make a difference!
Do you think this's plausible?
I always use PBS for washing cells. We have it made in house - you can buy PBS tablets if you don't want to make it up yourself. Make sure you sterilize it. Of course you can buy it already made but $$$.