{"doi":"10.1073/pnas.89.13.6020","title":"Detection of rare antigen-presenting cells by the lacZ T-cell activation assay suggests an expression cloning strategy for T-cell antigens.","abstract":"<jats:p>The alpha/beta T-cell receptor a complex ligand formed by the association of antigenic peptides with molecules of the major histocompatibility complex (MHC). The inherent limitations of the conventional T-cell activation assays used to detect these peptide/MHC ligands have, until now, hampered the development of expression cloning systems for T-cell antigens. To overcome these limitations, we have recently introduced a method for detecting ligand-induced activation of individual T cells. This assay, which makes use of a lacZ reporter construct, differs from conventional ligand-induced activation assays in that it allows the detection of single, activated T cells in large pools of resting cells. We applied the lacZ assay to the problem of screening expression libraries, which requires the ability to detect ligand-bearing antigen-presenting cells when they are present at very low frequency. We show here that ligand-expressing antigen-presenting cells can be detected at frequencies of 1:10(3)-10(4), a level of sensitivity compatible with the screening of cDNA libraries. Furthermore, by using as antigen-presenting cells COS-7 cells stably transfected with the murine Kb class I MHC molecule, we demonstrate that transiently expressed ovalbumin is efficiently processed and presented to an ovalbumin/Kb-specific T-cell hybridoma. lacZ expression is induced in a detectable number of cocultured T cells, even when the ovalbumin cDNA consists of only 1:10(4) of the total DNA used to transfect the COS cells. These results suggest that unknown T-cell antigens may be identified by screening cDNA libraries in MHC-expressing COS cells using lacZ-inducible T cells as indicators of peptide antigen expression.</jats:p>","journal":"Proceedings of the National Academy of Sciences","year":1992,"id":621164,"datarank":0.906750797105402,"base_score":6.045005314036012,"endowment":6.045005314036012,"self_citation_contribution":0.906750797105402,"citation_network_contribution":0.0,"self_endowment_contribution":0.906750797105402,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":421,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":null,"is_data_producer":false,"deposit_databanks":null,"is_oa":false,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":null,"fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1603811,"name":"S Sanderson","orcid":null,"position":1,"is_corresponding":false},{"id":1603813,"name":"N Shastri","orcid":null,"position":2,"is_corresponding":false},{"id":1603810,"name":"J Karttunen","orcid":null,"position":0,"is_corresponding":false}],"reference_count":0,"raw_metadata":{"has_enrichment":true,"resolved":true,"title":"Detection of rare antigen-presenting cells by the lacZ T-cell activation assay suggests an expression cloning strategy for T-cell antigens.","abstract":"<jats:p>The alpha/beta T-cell receptor a complex ligand formed by the association of antigenic peptides with molecules of the major histocompatibility complex (MHC). The inherent limitations of the conventional T-cell activation assays used to detect these peptide/MHC ligands have, until now, hampered the development of expression cloning systems for T-cell antigens. To overcome these limitations, we have recently introduced a method for detecting ligand-induced activation of individual T cells. This assay, which makes use of a lacZ reporter construct, differs from conventional ligand-induced activation assays in that it allows the detection of single, activated T cells in large pools of resting cells. We applied the lacZ assay to the problem of screening expression libraries, which requires the ability to detect ligand-bearing antigen-presenting cells when they are present at very low frequency. We show here that ligand-expressing antigen-presenting cells can be detected at frequencies of 1:10(3)-10(4), a level of sensitivity compatible with the screening of cDNA libraries. Furthermore, by using as antigen-presenting cells COS-7 cells stably transfected with the murine Kb class I MHC molecule, we demonstrate that transiently expressed ovalbumin is efficiently processed and presented to an ovalbumin/Kb-specific T-cell hybridoma. lacZ expression is induced in a detectable number of cocultured T cells, even when the ovalbumin cDNA consists of only 1:10(4) of the total DNA used to transfect the COS cells. These results suggest that unknown T-cell antigens may be identified by screening cDNA libraries in MHC-expressing COS cells using lacZ-inducible T cells as indicators of peptide antigen expression.</jats:p>","is_dataset_classified":null,"base_score":6.045005314036012,"endowment":6.045005314036012,"datacite_reuse_total":0,"file_count":0,"downloads":0,"views":0,"has_version_chain":false,"is_dataset":false,"is_oa":false,"pmid":"1378619","pmcid":"PMC402130","openalex_id":"https://openalex.org/W2048784307","authors":[],"funders":[{"funder_name":"NIAID NIH HHS","grant_id":"AI-26604","title":null}],"total_grants":1,"fwci":1.7559,"citation_percentile":0.85287638,"influential_citations":0,"citation_trend":[{"year":2012,"count":18},{"year":2013,"count":10},{"year":2014,"count":17},{"year":2015,"count":11},{"year":2016,"count":16},{"year":2017,"count":14},{"year":2018,"count":14},{"year":2019,"count":15},{"year":2020,"count":17},{"year":2021,"count":12},{"year":2022,"count":10},{"year":2023,"count":10},{"year":2024,"count":12},{"year":2025,"count":10},{"year":2026,"count":11}],"oa_status":"green","license":null,"oa_locations":[{"url":"https://pnas.org/doi/pdf/10.1073/pnas.89.13.6020","host_type":"publisher"},{"url":"https://doi.org/10.1073/pnas.89.13.6020","host_type":"journal"},{"url":"https://pubmed.ncbi.nlm.nih.gov/1378619","host_type":"repository"},{"url":"http://europepmc.org/articles/PMC402130","host_type":"repository"},{"url":"https://www.ncbi.nlm.nih.gov/pmc/articles/402130","host_type":"repository"}],"fields_of_study":["T-cell and B-cell Immunology","CAR-T cell therapy research","Immunotherapy and Immune Responses","Animals","Antigen-Presenting Cells","Biological Assay","Cell Line","Cloning, Molecular","DNA","Dose-Response Relationship, Immunologic","H-2 Antigens","Hybridomas","In Vitro Techniques","Interleukin-2","Ligands","Lymphocyte Activation","Mice","Ovalbumin","Staining and Labeling","T-Lymphocytes","beta-Galactosidase"],"mesh_terms":["Animals","Antigen-Presenting Cells","beta-Galactosidase","Biological Assay","Cell Line","Cloning, Molecular","DNA","Dose-Response Relationship, Immunologic","H-2 Antigens","Hybridomas","Interleukin-2","Ligands","Lymphocyte Activation","Ovalbumin","Staining and Labeling","T-Lymphocytes","Mice","In Vitro Techniques"],"keywords":["Antigen","Molecular biology","Major histocompatibility complex","Biology","Transfection","T cell","Complementary DNA","Ovalbumin","Epitope","Antigen-presenting cell","cDNA library","Cell culture","Immune system","Gene","Immunology","Genetics"],"sdg_mappings":[],"linked_datasets":[],"clinical_trials":[],"software_tools":[],"database_accessions":[],"source":"live","citation_network_status":"fetched"},"created_at":"2026-08-03T13:29:40.211879Z","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":[]}