Title
Measurement of Peptide Binding to MHC Class II Molecules by Fluorescence Polarization
Academic Program
Immunology & Microbiology
UMMS Affiliation
Department of Pathology; Department of Biochemistry and Molecular Pharmacology
Publication Date
2014-08-01
Document Type
Article
Disciplines
Biochemistry | Immunology and Infectious Disease | Immunology of Infectious Disease
Abstract
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.
DOI of Published Version
10.1002/0471142735.im0510s106
Source
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
Journal/Book/Conference Title
Current protocols in immunology / edited by John E. Coligan ... [et al.]
Related Resources
PubMed ID
25081912
Repository Citation
Yin L, Stern LJ. (2014). Measurement of Peptide Binding to MHC Class II Molecules by Fluorescence Polarization. Morningside Graduate School of Biomedical Sciences Student Publications. https://doi.org/10.1002/0471142735.im0510s106. Retrieved from https://escholarship.umassmed.edu/gsbs_sp/1930