weird & curious problem with CHO cell culture - (Mar/25/2011 )

After multiple postings here and lots of troubleshooting, I kinda figured why my CHO cells were dying. My CHO cells have been transfected with myoglobin and they grow in a special Ham's F12 media with addition of geneticin, hemin, 10% FBS and 1% P/S

I split my cells today and by next day my media is completely green in color and all cells are floating

Contamination was obviously suspected although others in the lab working on different cell lines weren't having such issues.

A grad student who worked on these cell lines before tells me that these cells are very sensitive to overgrowth ... beyond 70% confluence - they'll die... and i'd always let them grow overconfluent. We finally figured - whenever i split cells, i seeded too many in one plate and they double in like 12 hrs... they'd overgrow and die by the next day. maybe these cells secreted some products... byproducts of their disintegration perhaps... which made the media change color like that. the media has otherwise only phenol red and green is not one of its color indicators. so maybe its just the media combining with some products of the dying cells that makes it look green...

you think this theory is plausible?

suhas

I think it is unlikely - unless the medium is yellow (from the overgrowth) and the cells are secreting something blue... which would have to be secreted at a very high rate to colour the medium after only 24 hours.

CHO cells can be a pain to work with.

That is my contribution to the discussion .

It is quite strange. I wouldnt think it would be media related. As mentioned above the media would turn yellow to indicate acidity which is due to a high cell number. Have you confirmed that it isnt a contaminant?

PDGF on Thu Mar 31 02:21:02 2011 said:

CHO cells can be a pain to work with.

That is my contribution to the discussion .