{"doi":"10.1101/2024.12.10.627643","title":"Deconvoluting TCR-dependent &amp; -independent activation is vital for reliable Ag-specific CD4 <sup>+</sup> T cell characterization by AIM assay","abstract":"Abstract AIM assays are thought to detect antigen (Ag)-specific T cell responses in an HLA- and cytokine-independent manner. Recent studies using AIM assays identified prominent Th17-like (CCR6 + ) CD4 + T cells and circulating follicular T helper cells (cTfh) in anti-viral contexts, but were not observed with peptide/HLA tetramer staining. We demonstrate that CD39 + Treg-like and CD26 hi Th22-like cells can be activated by cytokines in a TCR-independent manner during in vitro Ag stimulation, leading to non-specific upregulation of prototypical AIM readouts. Transcriptional analysis of memory CD4 + T cells that underwent TCR-dependent or -independent activation enabled discrimination of bona fide Ag-specific T cells from cytokine-activated Treg and Th22 cells. CXCR4 downregulation emerged as a hallmark of clonotypic expansion and TCR-dependent activation in memory CD4 + T cells and cTfh. Tracking tetramer-binding cells during re-stimulation showed that CXCR4 - CD137 + cells provide a more accurate measure of the Ag-specific population than standard AIM readouts. This modified assay excludes the predominately CCR6 + cytokine-activated T cells that contribute to an average 12-fold overestimation of the Ag-specific population. As AIM assays enable the rapid study of T cell responses against emerging pathogens, our findings provide a highly accurate approach to characterize genuine Ag-specific T cell responses that contribute to protective immunity.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2024,"id":489394,"datarank":0.16479184330021646,"base_score":1.0986122886681096,"endowment":1.0986122886681096,"self_citation_contribution":0.16479184330021646,"citation_network_contribution":0.0,"self_endowment_contribution":0.16479184330021646,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":2,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9625,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2024-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1335019,"name":"Lauren Burmas","orcid":null,"position":1,"is_corresponding":false},{"id":555595,"name":"Hyon‐Xhi Tan","orcid":"0000-0003-1064-3934","position":2,"is_corresponding":false},{"id":1334719,"name":"Mai-Chi Trieu","orcid":"0000-0001-9181-5843","position":3,"is_corresponding":false},{"id":654817,"name":"Hyun Jae Lee","orcid":"0000-0002-9204-6078","position":4,"is_corresponding":false},{"id":1334720,"name":"Daniel Rawlinson","orcid":"0000-0002-3122-6243","position":5,"is_corresponding":false},{"id":242747,"name":"Ashraful Haque","orcid":"0000-0003-2260-0026","position":6,"is_corresponding":false},{"id":108826,"name":"Stephen J. Kent","orcid":"0000-0002-8539-4891","position":7,"is_corresponding":false},{"id":108825,"name":"Adam K. Wheatley","orcid":"0000-0002-5593-9387","position":8,"is_corresponding":false},{"id":473627,"name":"Jennifer A. Juno","orcid":"0000-0002-9072-1017","position":9,"is_corresponding":false},{"id":1334718,"name":"Ming Z. M. Zheng","orcid":"0000-0002-5406-7425","position":0,"is_corresponding":true}],"reference_count":64,"raw_metadata":{"citation_network_status":"fetched"},"created_at":"2026-07-19T02:08:28.548530Z","pmid":null,"pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}