Distinguishing Signal From Noise in Immunopeptidome Studies of Limiting-Abundance Biological Samples: Peptides Presented by I-A^b in C57BL/6 Mouse Thymus.

ABSTRACT: Antigen presentation by MHC-II proteins in the thymus is central to selection of CD4 T cells, but analysis of the full repertoire of presented peptides responsible for positive and negative selection is complicated by the low abundance of antigen presenting cells. A key challenge in analysis of limiting abundance immunopeptidomes by mass spectrometry is distinguishing true MHC-binding peptides from co-eluting non-specifically bound peptides present in the mixture eluted from immunoaffinity-purified MHC molecules. Herein we tested several approaches to minimize the impact of non-specific background peptides, including analyzing eluates from isotype-control antibody-conjugated beads, considering only peptides present in nested sets, and using predicted binding motif analysis to identify core

SUBMITTER: Nanaware PP 

PROVIDER: S-EPMC8116589 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature