no growth in TC plate - (Jan/17/2006 )

Hello all,

I am going crazy to solve this problem over past one month. Please please give your suggestions for troubleshooting.

I am working on animal cell based assays for studying enzyme induction upon challenge with steroids and cytotoxicity profiles. I am using five different cell lines for three different assays. The assays have been standardised and routinely practised in our laboratory.

Suddenly for past one month, I am facing problem with cell growth in 96 well plates. Cells show poor attachment, aggregation and visible death upon 24 hour incubation in microplates. This is despite of the healthy growth in parent flask for which same batch of media, incubators are used. I am using Nunc TC plates and flasks. All the cell culture products (media and tissue culture vessels) have been successfully used in past. I have neither changed vendor nor cat. no.
To confirm my findings, I tried growing cells in greiner cellstar plates. but similar results obtained.

In summary, I have tested five cell lines (HTC, L929, Hep2, HeLa, MDCK) for their growth in 96 well plate (both Nunc and greiner cellstar). Cells in parent flasks appear healthy but no growth observed in microplates. Media, reagents and incubators used for microplates and flasks are same.

Please help me in troubleshooting.

[quote name='BioBabe' date='Jan 17 2006, 12:51 PM' post='37273']


Hi I have had similar problems.... I was originally culturing in 100ul and found increasing to 200ul improves cell viability; particularly in the middle of wells; maybe your cells are starving??

Hello once again,

I tried growing cells in 6 well, 24 well and 48 well plates as well. Cells appear healthy, attached to the bottom (about 30-40% confluency) after 6 hr incubation. But viability is lost after 24 hr incubation, cells detach and dead cells float in media. Cells grow heathily in T25 / T75 flasks.

Any suggestions....pls.

someone once said to check that your little pan of water is full...I have also experienced that one....I hope that is your trouble; it is an easy fix even if you do feel silly