Using Real Time PCR for Cell Viability
Posted 23 March 2009 - 07:02 AM
Does anybody have any experience and advice with this?
Posted 23 March 2009 - 10:19 PM
If the control gene is not going to be affected by RNAi experiments, I think its expression can tell you the viability of the cells.
Posted 24 March 2009 - 05:37 AM
Posted 24 March 2009 - 03:32 PM
Posted 24 March 2009 - 06:23 PM
Amplifying genomic region may not tell you cell viability because dead cells may also contain the template.
Posted 24 March 2009 - 08:06 PM
Thanks for your reply, I'll look into it. Perhaps GAPDH would be another option, also seeing we have the primer/probes for this gene. Alternatively, would amplifying a genomic region be another option, seeing that remains pretty constant in the cells?
GAPDH is a good candidate as an internal control for qRT-PCR. Trust me, 18S and 28S RNA would save you lots of time. My personal opinion is that results of qRT-PCR was too indirect to demostrate cell viability change, especially when cell viability is a key issue in your research. If you simply want to use the result as an implication, it would be fine. Actually, MTT is very cheap and more convincible. May it help you and good luck.
Posted 27 March 2009 - 04:32 AM
I am familiar with the MTT assay. I should have mentioned that I intend to perform cell viability assays for cells that have undergone knock-down or upregulation of genes by transfection. The reason I would like to choose TaqMan as a cell viability assay is because I believe I would waste more money, time and effort in performing two transfections in parallel (one for qRT-PCR and the other for cell viability) for an MTT assay than using the same cDNA for two different qRT-PCR assays.
As two members here recommend the 18S and 28S RNA as good candidates I will probably go by those.