I did SILAC experiment for a drug toxicity. Do I need to validate the data? As a procedure, SILAC is an established technique. So the data I got is good quality. If I have to do some validation work, then what will be preferable procedure?
Again, as the correlation between genomic data and proteomics data is very low, is it wise to go for global gene expression analysis? In comparison between genomic and proteomics, which one the best in molecular pathway analysis?
Is there any way to select proteins for validation or its a random selection? From my 101 proteins ( altered), I select 2 up and 2 down regulated protein randomly. Their fold changes were higher. But abundance of proteins were seems very low. So, in western blot, a faint signals were found. Those were as same as proteomic result, though so faint difference. Can you advise me?
Thank you very much. I chose proteins for validation based on their fold changes. I took highest fold changing proteins (3-4 folds). You are so correct. My WB results seems correct but differences between control and reactions are so small. It may be due to its low abundance. I checked with literature which showed abundance is as low as 0.005 microgram/micrllitre. Usually for WB, it is commendatory that protein of interest should be at least 5 microgram/lane. I got it only 0.125 microgram/lane. Even sensitivity of antibody is also a matter. It seems relatively low, as they recommend as 1:50 -100 dilution. So, ELISA may be the best way. I will run ELISA again. Thanks.
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