Needing a consistant cell suspension - Non-homogenous cells make me crazy! (Nov/10/2005 )

Hi All,

I am setting up a lot of antiproliferative assays with LnCAP cells in 96-well plates. I'm having a lot of trouble getting a consistant number of cells per well (aiming for 10000 per well, based on the hematacytometer count).

It seems that no matter how many times I pipette the mixture up and down, I still get an unacceptable level of variation between wells. It's not so bad for other cell cultures, but LnCAP seems particularly prone to clumping.

Does anyone have any suggestions on how to prevent this? Thanks.

Would it be possible to increase the volumes you are working with??

How are you detaching the cells initially, trypsin or other?? Some people will tap the plate to release cells when trypsinizing and that will cause more clumps than if you just wait until the cells come off by themselves...

Potentially trypsinizing longer will also help or adding more EDTA when detaching???

HTH--good luck

Thanks, I'll try a longer trypsin treatment when I plate out cells on Monday.