In Situ Peptide-MHC-II Tetramer Staining of Antigen-Specific CD4(+) T Cells in Tissues

The invention of peptide-MHC-tetramer technology to label antigen-specific T cells has led to an enhanced understanding of T lymphocyte biology. Here we describe the development of an in situ pMHC-II tetramer staining method to visualize antigen-specific CD4(+) T cells in tissues. This method complements other methods developed that similarly use MHC class II reagents to stain antigen-specific CD4(+) T cells in situ. In this study, we used group A streptococcus (GAS) expressing a surrogate peptide (2W) to inoculate C57BL/6 mice, and used fresh nasal-associated lymphoid tissues (NALT) in optimizing the in situ staining of 2W:I-A(b) specific CD4(+) T cells. The results showed 2W:I-A(b) tetramer-binding CD4(+) T cells in GAS-2W but not GAS infected mice. This method holds promise to be broadly applicable to study the localization, abundance, and phenotype of antigen-specific CD4(+) T cells in undisrupted tissues.