I would like to investigate how a kind of hormone influences a sort of gene`s expression in pancreatic cells( INS-1), and I am about to make a concentration gradient of this hormone now to determine an optimal concentration.
There are similar literature on apoptosis analyze of the same cell line by this hormone in which suggests a group of concentrations. But I am not sure if I can take these articles for reference or not, because what I would like to do is to see what is the potential involvement of signaling pathways when the gene expression is decreased by this hormone. I think for this we must keep the cell alive basically.
Maybe the question sounds stupid, I am still so confused about these concepts
・What is the relationship between apoptosis and gene expression ?
・How to make concentration gradient?any rules or protocols?
・In my case, is it ok for me to take their concentration (literature mentioned above) for reference?
Your kind reply will be greatly appreciated, thank you all.
As per my knowledge, the relation between apoptosis and gene expression would not be same for all genes.. Some genes get upregulaed and some downregulated depending upon the pathways they are involved in.. Usually the pro-apoptotic genes and the genes associated with cell death pathways will be upregulated and the anti-apoptotic and genes associated with cell survival pathways get downregulated.. Also some genes involved in inflammation initially get upregulated and then later downregulated (transient expression).. So this is variable..
To make a concentration gradient of the hormone.. you need a specific assay which can detect your hormone of interest.. ELISA is an option some even do RIA or other immune assays.. even HPLC in somecases... Ultimately you need to normalise it with the protein content in order to get the actual increase in concentration..
Its always better to make your own curve as sometimes handling, treatment, cell passage no. will affect such things..
Best..
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