Login

Join for Free!
117133 members


FACS and growth curve result

Discussion of all aspects of cellular structure, physiology and communication.

Moderator: BioTeam

FACS and growth curve result

Postby nusrat1977 » Fri Mar 25, 2011 1:39 am

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
nusrat1977
Garter
Garter
 
Posts: 2
Joined: Fri Mar 25, 2011 1:32 am

Postby canalon » Fri Mar 25, 2011 3:18 am

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.
Patrick

Science has proof without any certainty. Creationists have certainty without
any proof. (Ashley Montague)
User avatar
canalon
Inland Taipan
Inland Taipan
 
Posts: 3909
Joined: Thu Feb 03, 2005 2:46 pm
Location: Canada

Postby nusrat1977 » Fri Mar 25, 2011 5:14 am

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.
nusrat1977
Garter
Garter
 
Posts: 2
Joined: Fri Mar 25, 2011 1:32 am


Postby canalon » Fri Mar 25, 2011 1:50 pm

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.
Patrick

Science has proof without any certainty. Creationists have certainty without
any proof. (Ashley Montague)
User avatar
canalon
Inland Taipan
Inland Taipan
 
Posts: 3909
Joined: Thu Feb 03, 2005 2:46 pm
Location: Canada

Re: FACS and growth curve result

Postby mohit » Sat Mar 26, 2011 5:09 am

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.
Last edited by JackBean on Sun May 22, 2011 4:34 pm, edited 1 time in total.
Reason: link removed
mohit
Garter
Garter
 
Posts: 4
Joined: Sat Mar 26, 2011 5:03 am


Return to Cell Biology

Who is online

Users browsing this forum: No registered users and 1 guest

cron