{"doi":"10.1016/j.xpro.2024.103310","title":"Protocol to quantify the activation dynamics of tumor-associated T cells in mice by functional intravital microscopy","abstract":"Tumor-associated T cells orchestrate cancer rejection after checkpoint blockade immunotherapy. T cell function depends on dynamic antigen recognition through the T cell receptor (TCR) resulting in T cell activation. Here, we present an approach to quantify the dynamics and magnitude of tumor-associated T cell activation at multiple time points in living mice using the genetically encoded calcium reporter Salsa6f and functional intravital microscopy (F-IVM). Our protocol allows researchers to measure the activation dynamics of various immune cells in vivo . For complete details on the use and execution of this protocol, please refer to Geels et al. 1 • Longitudinal 2-photon intravital imaging protocol for T cell dynamics in a skin tumor • Quantification of TCR signaling using the ratiometric fluorescent sensor Salsa6f • Steps include surgery, image acquisition, and analysis using Imaris and MATLAB Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Tumor-associated T cells orchestrate cancer rejection after checkpoint blockade immunotherapy. T cell function depends on dynamic antigen recognition through the T cell receptor (TCR) resulting in T cell activation. Here, we present an approach to quantify the dynamics and magnitude of tumor-associated T cell activation at multiple time points in living mice using the genetically encoded calcium reporter Salsa6f and functional intravital microscopy (F-IVM). Our protocol allows researchers to measure the activation dynamics of various immune cells in vivo .","journal":"STAR Protocols","year":2024,"id":477571,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"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":0.9506,"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":1201376,"name":"Claire Murat","orcid":"0000-0003-2359-6444","position":1,"is_corresponding":false},{"id":492643,"name":"Alexander Moshensky","orcid":"0000-0002-1778-4601","position":2,"is_corresponding":false},{"id":417469,"name":"Shivashankar Othy","orcid":"0000-0001-6832-5547","position":3,"is_corresponding":false},{"id":356716,"name":"Francesco Marangoni","orcid":"0000-0002-2490-849X","position":4,"is_corresponding":false},{"id":623867,"name":"Shannon N Geels","orcid":"0000-0002-7991-6104","position":0,"is_corresponding":true}],"reference_count":27,"raw_metadata":null,"created_at":"2026-07-19T02:06:37.812633Z","pmid":"39306849","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":[]}