I have some active compounds and I want to check the effect of those compounds on p38 MAPK pathway genes (p38, JNK and ERK1/2) or may by AKT/PKB and PTEN in MCF7 and MDA-MB-231 breast cancer cell lines. I cannot go for western blot or immunocytochemistry due to lack of antibodies. Is it OK to check effect of active compounds on just gene expression and not on protein expression in case of above mentioned pathways?
I recommend the PCR-Array from Qiagen:
https://www.qiagen.com/us/shop/pcr/real-time-pcr-enzymes-and-kits/two-step-qrt-pcr/rt2-profiler-pcr-arrays/#products97eefe50-7f5e-4918-96a1-02235877cdd8
its ok to check gene expression for these by qPCR, but I'm not sure if you will see much effect by any compound (unless it interferes with transcription somehow). Generally, effects on these kinases have to do with their phosphorylation states rather than their expression. For that you would need the phospho- antibodies. You could also preform kinase assays to look at their activity.
Hi Rehan
It is okay, yes, PCR is a very reliable technique to check gene expression changes due to treatments. However, mRNA switches on and off depending on several reasons, so if you see a downregulation of mRNA (meaning it is not significant vs your controls) it does not necessarily mean your cellular function or protein trnaslation is not there neither. it also depends on what you want to show
i worked with P38 before and I used this antibody https://www.cellsignal.com/products/primary-antibodies/p38-mapk-antibody/9212 and worked fine for me
good luck!
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