Patent ID: 9110080
Filing Date: 2015-08-18
Classification: G01N

Abstract:
1. A method of assaying the affinity of a peptide of interest for binding to an individual MHC class I molecule, the method comprising the steps of: labeling a first peptide with a fluorescent label, wherein the first peptide is known to bind to a specific MHC class I molecule; activating individual soluble, specific MHC class I molecules by heating the individual soluble, specific MHC class I molecules for about 15 minutes at about 53° C. to cause the individual soluble, specific MHC class I molecules to release any endogenous peptide loaded therein, thereby activating the individual soluble, specific MHC class I molecules; mixing the labeled first peptide and the activated individual soluble, specific MHC class I molecules together with different concentrations of a second peptide, wherein the second peptide is a peptide of interest, to provide individual soluble, specific MHC class I molecule-labeled first peptide complexes and individual soluble, specific MHC class I molecule-second peptide complexes, wherein the individual soluble, specific MHC class I molecule-first peptide complexes are detectable by fluorescence polarization; and determining a binding affinity of the peptide of interest for the individual soluble, specific MHC class I molecule by determining an inhibitory concentration of the second peptide that inhibits 50% binding of the first peptide to the individual soluble, specific MHC class I molecule.