{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T06:01:46Z","timestamp":1785391306510,"version":"3.55.0"},"reference-count":64,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T00:00:00Z","timestamp":1785369600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Immunol."],"abstract":"<jats:p>\n                    Tumor-associated B cell responses are frequently observed in tumors and their draining lymph nodes, yet their antigen specificities and functional potential remain incompletely defined. Historically, intracellular antigens have been largely disregarded in tumor antigen mining and discovery efforts. In particular, the extent to which intracellular tumor antigens can be effectively targeted by antibodies and the cellular immune system has remained unclear, poorly characterized, and therapeutically underexploited. We developed a high-throughput platform combining hybridoma generation with unbiased antigen-based flow cytometric enrichment to isolate tumor-reactive monoclonal antibodies. Using a murine 4T1 triple-negative breast cancer model engineered to express the intracellular protein ZsGreen1 (ZsG), we characterized B cell responses and selected antigen-specific hybridomas via fluorescence-activated cell sorting. Antibody specificity was validated using biochemical and cellular assays, and\n                    <jats:italic>in vivo<\/jats:italic>\n                    activity was evaluated following systemic administration. Translational applicability was assessed using hybridomas generated from human tonsillar and peripheral blood B cells. Tumor growth elicited a class-switched, antigen-specific B cell response against the intracellular ZsG antigen. Hybridomas retained a dual phenotype, expressing both surface B cell receptors and secreted immunoglobulins, enabling efficient antigen-driven selection. Isolated monoclonal antibodies recognized native intracellular epitopes that were not detectable using conventional screening approaches. Notably, systemic administration of an anti-ZsG antibody was associated with reduced tumor growth in early established tumors\n                    <jats:italic>in vivo<\/jats:italic>\n                    , supporting functional antibody activity under these conditions. Furthermore, biotinylated tumor lysates enabled unbiased identification of additional tumor-reactive antibodies, and the platform was successfully extended to human B cells. These findings support the accessibility of intracellular tumor antigens to antibody recognition and establish a framework for discovering antibodies targeting this antigen class. The platform described here provides a scalable approach for isolating tumor-reactive antibodies and expands the landscape of potential targets for cancer immunology and immunotherapy.\n                  <\/jats:p>","DOI":"10.3389\/fimmu.2026.1867422","type":"journal-article","created":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T05:40:38Z","timestamp":1785390038000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":0,"title":["B cell receptor\u2013based hybridoma enrichment enables discovery of antibodies against intracellular tumor antigens"],"prefix":"10.3389","volume":"17","author":[{"given":"Iris","family":"Dotan","sequence":"first","affiliation":[{"name":"School of Neurobiology, Biochemistry and Biophysics, The George S. Wise Faculty of Life Sciences, Tel Aviv University","place":["Tel Aviv, Israel"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yuval","family":"Raviv","sequence":"additional","affiliation":[{"name":"School of Neurobiology, Biochemistry and Biophysics, The George S. Wise Faculty of Life Sciences, Tel Aviv University","place":["Tel Aviv, Israel"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"D\u00e9bora R.","family":"Bublik","sequence":"additional","affiliation":[{"name":"School of Neurobiology, Biochemistry and Biophysics, The George S. Wise Faculty of Life Sciences, Tel Aviv University","place":["Tel Aviv, Israel"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jule A.","family":"Kortendieck","sequence":"additional","affiliation":[{"name":"School of Neurobiology, Biochemistry and Biophysics, The George S. Wise Faculty of Life Sciences, Tel Aviv University","place":["Tel Aviv, Israel"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shrey","family":"Jain","sequence":"additional","affiliation":[{"name":"School of Neurobiology, Biochemistry and Biophysics, The George S. Wise Faculty of Life Sciences, Tel Aviv University","place":["Tel Aviv, Israel"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yifat","family":"Glucksam Galnoy","sequence":"additional","affiliation":[{"name":"School of Neurobiology, Biochemistry and Biophysics, The George S. Wise Faculty of Life Sciences, Tel Aviv University","place":["Tel Aviv, Israel"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nechama I.","family":"Smorodinsky","sequence":"additional","affiliation":[{"name":"The Shmunis School of Biomedicine and Cancer Research, The George S. 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