biostudies-literatureUnknown8(3)McComb S<h4>Background</h4>Production of chimeric antigen receptor T cell (CAR-T) therapies depends on antibody reagents to label, isolate, and expand T cell products. We sought to create antibody tools specific for the variable domain of heavy-chain only antibodies (VHHs), also known as nanobodies, used in some CARs.<h4>Methods</h4>We immunized a mouse with VHH and selected two murine monoclonal antibodies (mAbs) that bind to distinct epitopes in conserved framework regions of llama-derived VHHs, and not to human VH domains. Anti-VHH mAbs were characterized by enzyme-linked immunosorbent assay, surface plasmon resonance, and hydrogen-deuterium exchange mass spectrometry; were then tested for cell/tissue labeling and for modulating cellular activity in VHH-CAR-T cells.<h4>Results</h4>We produced aAntibody therapeutics242-258https://www.ebi.ac.uk/biostudies/studies/S-EPMC12451263biostudies-literatureBroadly reactive anti-VHH antibodies for characterizing, blocking, or activating nanobody-based CAR-T cells.PMC12451263Aubry AAcchione MCoutu MAcel AWu CMarcil AJoubert STremblay TLMoraitis ANParat MHill JJZafer AWeeratna RDMalenfant FPon RAPohankova PPage MShepherd AHussack GEl Bakkouri MLippens JRaphael SPlante PBennychen BSheff JLamoureux LNguyen TArbabi-Ghahroudi MWebb JSmith EMcComb SDupont BFortin AGadoury CTamblyn LFaulkes SManceur APSchrag JZhu QfalseBroadly reactive anti-VHH antibodies for characterizing, blocking, or activating nanobody-based CAR-T cells.<h4>Background</h4>Production of chimeric antigen receptor T cell (CAR-T) therapies depends on antibody reagents to label, isolate, and expand T cell products. We sought to create antibody tools specific for the variable domain of heavy-chain only antibodies (VHHs), also known as nanobodies, used in some CARs.<h4>Methods</h4>We immunized a mouse with VHH and selected two murine monoclonal antibodies (mAbs) that bind to distinct epitopes in conserved framework regions of llama-derived VHHs, and not to human VH domains. Anti-VHH mAbs were characterized by enzyme-linked immunosorbent assay, surface plasmon resonance, and hydrogen-deuterium exchange mass spectrometry; were then tested for cell/tissue labeling and for modulating cellular activity in VHH-CAR-T cells.<h4>Results</h4>We produced a2025-01-01T00:00:00Z2025 Jul2026-06-03T19:19:11.379Z2026-05-30T03:06:49.475ZS-EPMC124512634098910610.1093/abt/tbaf011