unwanted cell clumping after splitting MCF7 cells - (Sep/11/2006 )

Hi there,

after splitting my MCF7 cells and spinning them down, I end up with a clump of cells that is really hard to resuspend and I would like to a single cell suspension for FACS purposes. I use the standard trypsin/EDTA .25% for splitting, incubation time is 5-10 min. Has anyone experience with that?

Thanks a bunch,

-S

You probably over treated cells and some of them lysed and released DNA. DNA tangled the remaining cells together.

if you didn't, you'll want to remove the trypsin from the cells and suspend in your basal media without FBS present. When resuspending your cell pellot you'll need to lightly triturate the cells with your pipette.

for my MCF-7s, i usually trypsinize for about 5-7 mins then add my media. pipette up and down a couple times then run it through a sterile strainer a couple times as well and theyre pretty good to go.

i trypsinise until they come off the plate, add media, spin, resuspend, and run it through an 18 gage syringe 5 times.

V