I am analyzing spleen, lymph nodes, and PBMCs for antigen specific CD8+ T cells. I want to gate out B cells, CD4+ T cells, monocytes, and NK cells (put all markers into a dump gate FL channel). Would CD19, CD4, CD11b, and CD335 work for this purpose?
Never used anti-CD335 but that with the other antibodies should work
Dear Jim: This review may be useful to you
ProtocolAntigen Processing
Volume 960 of the series Methods in Molecular Biology pp 309-326 Tracking Antigen-Specific CD8+ T Cells Using MHC Class I Multimers
Cécile Alanio Isabelle Bouvier, Hélène Jusforgues-Saklani, Matthew L. Albert
The tracking of epitope-specific T cells is a useful approach for the study of adaptive immune responses. This protocol describes how Major Histocompatibility Complex Class I (MHC-I) multimers can be used to stain, enrich, and enumerate (rare) populations of CD8+ T cells specific for a given antigen. It provides the detailed steps for multimer labeling, magnetic enrichment, and cytometric analysis. Additionally, it provides informations for multiplexing experiments in order to achieve simultaneous detection of multiple antigenic specificities, and strategies for coupling the protocol with functional assays (e.g., intracellular cytokine staining). Future developments in cytometric systems (e.g., mass spectroscopy-based cytometry) and gene expression studies (e.g., single cell PCR) will extend these approaches and provide an unprecedented assessment of the immune repertoire.
Key words
CD8+ T cells MHC Class I multimers Antigen-specific T cells T-cell receptor
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