Measurement of Peptide Binding to MHC Class II Molecules by Fluorescence Polarization
Immunology & Microbiology
Department of Pathology; Department of Biochemistry and Molecular Pharmacology
Biochemistry | Immunology and Infectious Disease | Immunology of Infectious Disease
Peptide binding to major histocompatibility complex class II (MHCII) molecules is a key process in antigen presentation and CD4+ T cell epitope selection. This unit describes a fairly simple but powerful fluorescence polarization-based binding competition assay to measure peptide binding to soluble recombinant MHCII molecules. The binding of a peptide of interest to MHCII molecules is assessed based on its ability to inhibit the binding of a fluorescence-labeled probe peptide, with the strength of binding characterized as IC50 (concentration required for 50% inhibition of probe peptide binding). Data analysis related to this method is discussed. In addition, this unit includes a support protocol for fluorescence labeling peptide using an amine-reactive probe. The advantage of this protocol is that it allows simple, fast, and high-throughput measurements of binding for a large set of peptides to MHCII molecules.
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Citation: Curr Protoc Immunol. 2014 Aug 1;106:5.10.1-5.10.12. doi: 10.1002/0471142735.im0510s106. Link to article on publisher's site
Current protocols in immunology / edited by John E. Coligan ... [et al.]
Yin, Liusong and Stern, Lawrence J., "Measurement of Peptide Binding to MHC Class II Molecules by Fluorescence Polarization" (2014). GSBS Student Publications. 1930.