Characterizing the specificity of T follicular helper cell repertoire in human peripheral blood and tonsil samples

T follicular helper cells (Tfh) are specialized CD4+ T cells, primarily localized in germinal centers (GCs) in secondary lymphoid organs. These cells play a key role in supporting B cell responses and selection of affinity maturated antibodies. The majority of research to date on human Tfh cells has been restricted to circulating Tfh-like cells (cTfh) from peripheral blood because of difficulties in obtaining secondary lymphoid organs. However, the relationship between Tfh cells in lymph nodes (follicular Tfh) and cTfh cells as well as their distinctiveness from the CXCR5â memory CD4+ T cell compartment are still unclear. By using donor-matched blood and tonsil samples, this project aimed to investigate the relationship between peripheral and follicular Tfh cells and also between Tfh and non-Tfh cell populations. Phenotypic analyses characterized and identified populations of blood (Tfh-Th1, Tfh-Th2 and ROM) and tonsil (Tfh-GC, Tfh-Th1, Tfh-Th2 and ROM) cells to be compared. Targeted sequence-based analysis of the T cell receptor (TCR) showed a repertoire overlap between peripheral Tfh subsets and tonsillar Tfh cells as well as the distinction between Tfh and the rest of CD4+ memory T cells. This was also confirmed in an antigen-specific system by comparing the peptide specificity and TCR repertoires of influenza virus haemagglutinin (HA) responsive Tfh and non-Tfh cells. Tfh and non-Tfh cell clones were also generated and their avidity characterized for use in Tfh and B cell co-cultures assays and for high resolution investigation of molecule trafficking in immune synapses. Correlation between peripheral and tonsillar compartments as well as distinction between Tfh and non-Tfh cells will inform the evaluation of Tfh responses in future clinical studies.