FACS and growth curve result

[nusrat1977]

Hello,
I am a chemist, do not know much about cell biology. Recently I have started some work. I took a cell line which express GFP. I studied the cell growth under various conditions and observed the number of cells are growing more in some cases. Then I did FACS. But I observed FACS result is same for all cases. Do FACS result has to corroborate growth curve? I thought FACS gives the % of cells expressing GFP which should be same irrespective of the number of cells. Am I wrong?
Please reply.
Thanks
Nusrat

[canalon]

It your cells are expressing the GFP constitutively then you would expect your %age to be constant. But I thought you could also get the number of cells in the sample. So if you have standardized your samples (same volume or something) *that* should corroborate your growth curves.

[nusrat1977]

Thank you very much for your reply. you are right, for standardized samples it should match the growth curves. But I remembered one thing. It also depends on the time factor i.e if the cells are dead, do the intensity of already expressed GFP dies to zero? I have to figure out.

[canalon]

There are quite a few version of the GFP as far as I know. The original gene/protein is not very stable IIRC, and more stable versions have been developed. And also some less stable too, because sometimes you want the protein to disapear quickly.
One way to test the stability of your gene in your cell line would be to have it expressed under a regulated promoter, induce for a while and then remove the inductor and make a time course measurement of the fluorescence intensity.

[mohit]

While studying the growth curve one should also be very aware about the different terminologies which are actually being employed. These may be Idiophase, trophophase. One should also consider the concept of biphasic growth which is also kknown as di auxic growth.