Why won't you grow for me?
October 1, 2008 6:59 AM Subscribe
Specific Cell culture question:
posted by LunaticFringe to Science & Nature (10 answers total)
I'm currently trying to culture the Dempsey cell line (ATCC# CCL-28) and am having no luck. The cells are just not replicating. They're alive but they don't repicate at all. We've double and triple checked the conditions described on the ATCC page and according to what's out there we should be golden. We've also checked a few lab pages and the references and everything matches what we're doing. Has anyone worked with these cells? I feel like there's something extremely obvious that I'm missing.
Here is our exact culture conditions:
McCoy's 5a Media from Gibco (or whatever company Gibco is now) supplemented with L-glutamine (1.5 mM) and sodium bicarb (2.2g/L) and 20% FBS (heat inactivated - is this the problem?)
37 deg C and 5% CO2
Cultured in 10 cm plates (Sarstedt) - uncoated (are we supposed to coat them with something?)
The L-glutamine is frozen in one-shot tubes and our other lines grow just fine, including a couple MES lines, so I don't think it's bad glutamine. The bicarb is culture-grade powder and, again, the other lines growing are just happy as can be so I don't think it could be that. We have no tested for mycoplasma but a test kit is on the way, just in case. Checking via DAPI has revealed no contamination but we're gonna do the PCR-based just to be extra sure. We haven't had any mycoplasma problems before but we've just received three new lines so I think a test would be prudent even if we weren't having issues with this line.
Thanks for the help everyone and sorry for the length.