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[MHCetc]

Hi, everybody. I do a lot of competition binding experiments with peptides and HLA. I'm in the process of optimizing the conditions so the IC50 I get is approximately what the Kd would be in a direct binding experiment. Probably I'm misunderstanding something here, but what does the Kd tell you about a ligand if you don't have information about the assay conditions? Is it simply convention that most labs tend to work in the same range?

Let me give an example. If somebody says the interaction between ligand A and receptor B has an IC50 of 3 nM, that doesn't tell you much unless you know a.) what the concentration of the target receptor is, b.) what the concentration of the labeled agonist is, and c.) how high the affinity of the labeled agonist is. It just says that at 3 nM, half of the agonist's specific binding has been inhibited by the test ligand. If the ligand was added at 0.1 pM, that's not saying much. Similarly, if you tweak the assay parameters so that 3 nM is a reasonable approximation of the Kd, there is still no information without knowing the assay parameters; i.e. at 3 nM, the test ligand saturates half of the available specific binding sites, but half of how much?

This is why I don't understand why websites like the Immune Epitope Database (IEDB) provide an enormous compendium of MHC binding assay results from different labs as if they were prima facie comparable. Does anybody have any insight on this matter?

Thanks,

Ryan

Edited by MHCetc, 21 July 2011 - 07:28 AM.