Hello all,
I am planning to perform a viability drug screen with Selleckchem bioactive compounds library (~2000 drugs).
My goal is to find drugs that differentially kill cells with wt or knockout of my gene of interest. Since the potency of drugs varies widely (nM to >50 uM range of IC50) I wonder what is the best design strategy. I saw that people just use 1 uM and/or 10 uM in primary screens but this would miss the relevant range of many drugs, where I have a chance to see the effect of my gene.
Obviously, doing the screen with more concentrations will decrease the replicates for each (I am thinking to use duplicates since CellTiter-GLo gives very tight replicates).
I would appreciate any input regarding the screen design.
Thanks,
Igor
Dear Igor,
I'm not really an expert, but did some more or less extensive screening approaches with some assay setups. I agree, that a fixed "high" concentration in the beginning for all compound would be the best strategy, but in you case the very high concnetrations would produce additional problems.
In the range of 100µM you will probably find a lot of toxic compounds, that generally will kill cells....any kind.
So you detect toxicity for wt and ko, without knowing, if you would get a more differentiated picture in the lower range.
I think you will probably need a more sophisticated guess, depending upon your target.
When I got it right, the "Selleckchem bioactive compounds library" appears to be a predesigned product.
You might contact Selleckchem and ask them for a general Tox profile for these compounds. Maybe they have somthing like that for several often used cell lines, maybe also for your wildtype cell line?!
Than you could choose individually, compound by compound. More work, but less painfull when you try to interpret your results.
We do automated high throughput screening in our facility for Chemical Genomics. For an initial screen, all compounds are used at 10 uM in DMSO. If you identify a hit, we then do a dose response curve over a broader range from 1 nM to 100 uM.
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