2 replies to this topic

[steve-irl]

Hey,

Am culturing HK-2 cells and am trying to detach them without using trypsin as it'll cleave a cell surface protein I want to look at by flow cytometry. Any ideas? I've tried using various concentrations of EDTA in PBS...the cells round up but won't come off. Am I not using a high enough concentration (I've tried 50mM) or am I just not leaving it on long enough. Any help GREATLY appreciated!

[wirly]

Maybe you could try the Cell Disociation buffers offered by Gibco/Invitrogen.  
https://catalog.invi...83&npc=2&nc=79
They did not work much better than EDTA for me but they may on your cell line.
For how long are you incubating with EDTA?  If it's a hearty cell they can take long exposures without dying.  Do you try to actively wash them off the plate using a smaller bore pipette?  I have not use HK-2 cell before, they may be like MDCKs which HATE to come off the plate w/o trypsin.

Good luck!

[anilkumarpr]

Hi

There are temperature sensitive surfaces for cell culture.  Keep cells cultured on temp sensitive surface at room temp for a few minutes and cells are detached as sheets.

Amazing isnt it

Try search on PIPAAm grafted surfaces for cell culture

Anil