I know that it depends on many things, such as the mechanism, but I was wondering about an average in order to plan stimulus time. I'm plannig to stimulate podocyte cells with some factors, such as TNF-A, IL1b, VEGF, sFlt1 and human sera. In our laboratory we have stimulated similar cells with human sera in the order of hours (12,24,48 h), but mainly to check at the protein level. In some literature I've read, in similar cells (glomerular endothelium), stimulus effects with TNF-A were best at 1 or 2 hours, and tend to disappear at 12 or 24 h. I know that this depends on the experiments own nature, but, which stimulus time upper and lower limits should I consider?
Based on my experience with murine astroglia stimulated with IL-6 is to look at an early (90-120min) and late (12-24hr). I believe this will give you a sufficiently complete picture. For global gene expression these are good starting points, but if you are going to look at specific genes, probably best to first look at many time points (30min-24hr) and use qPCR or RPA to determine the kinetics and use the most optimal time point for future experiments. Of course, if this information is already available in the literature then half the work is done for you already. Best of luck.
Thanks, Ricardo, I also think it's better to test the kinetics for each case, but ,unfortunately, in this case, I can't, since I have RT-qPCR arrays with primers attached, so I can't perform previous experiments. I think I'll have to rely on literature.
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