title: Efficient ex vivo analysis of CD4+ T-cell responses using combinatorial HLA class II tetramer staining.
abstract: MHC tetramers are an essential tool for characterizing antigen-specific CD4+ T cells. However, their ex vivo analysis is limited by the large sample requirements. Here we demonstrate a combinatorial staining approach that allows simultaneous characterization of multiple specificities to address this challenge. As proof of principle, we analyse CD4+ T-cell responses to the seasonal influenza vaccine, establishing a frequency hierarchy and examining differences in memory and activation status, lineage commitment and cytokine expression. We also observe cross-reactivity between an established epitope and recent variant and provide a means for probing T-cell receptor cross-reactivity. Using cord blood samples, we correlate the adult frequency hierarchy with the naive precursor frequencies. Last, we use our combinatorial staining approach to demonstrate that rheumatoid arthritis patients on therapy can mount effective responses to influenza vaccination. Together, these results demonstrate the utility of combinatorial tetramer staining and suggest that this approach may have broad applicability in human health and disease.
authors: Benaroya Research Institute at Virginia Mason, Translational Research Program, Seattle, Washington 98101, USA.|Neuroimmunology Unit, Department of Neuroscience, Solna, Karolinska Institutet, Karolinska University Hospital, SE-171 76 Stockholm, Sweden.
authors: Hannes
authors: H
authors: Uchtenhagen
authors: Benaroya Research Institute at Virginia Mason, Translational Research Program, Seattle, Washington 98101, USA.
authors: Cliff
authors: C
authors: Rims
authors: Benaroya Research Institute at Virginia Mason, Diabetes Program, Seattle, Washington 98101, USA.
authors: Gabriele
authors: G
authors: Blahnik
authors: Benaroya Research Institute at Virginia Mason, Diabetes Program, Seattle, Washington 98101, USA.
authors: I-Ting
authors: IT
authors: Chow
authors: Benaroya Research Institute at Virginia Mason, Diabetes Program, Seattle, Washington 98101, USA.|Department of Medicine, University of Washington, Seattle, Washington 98195, USA.
authors: William W
authors: WW
authors: Kwok
authors: Benaroya Research Institute at Virginia Mason, Translational Research Program, Seattle, Washington 98101, USA.
authors: Jane H
authors: JH
authors: Buckner
authors: Benaroya Research Institute at Virginia Mason, Diabetes Program and Tetramer Core Laboratory, Seattle, Washington 98101, USA.
authors: Eddie A
authors: EA
authors: James
mesh_terms: D000328:Adult; D000368:Aged; D001172:Arthritis, Rheumatoid / Q000097:blood / Q000188:drug therapy / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D003429:Cross Reactions; D018984:Epitopes, T-Lymphocyte / Q000276:immunology*; D005260:Female; D005312:Fetal Blood; D059811:HLA-DRB1 Chains / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D007252:Influenza Vaccines / Q000276:immunology; D007251:Influenza, Human / Q000517:prevention & control; D008213:Lymphocyte Activation / Q000276:immunology*; D008297:Male; D008875:Middle Aged; D055503:Protein Multimerization; D011948:Receptors, Antigen, T-Cell / Q000276:immunology; D013194:Staining and Labeling / Q000379:methods*
chemical_list: D018984:Epitopes, T-Lymphocyte; D059811:HLA-DRB1 Chains; D007252:Influenza Vaccines; D011948:Receptors, Antigen, T-Cell