biostudies-literatureUnknown11(4)Hu NBotulinum neurotoxin (BoNT) is a highly lethal toxin produced by the anaerobic bacterium Clostridium botulinum, which leads to nerve paralysis following poisoning. At present, there is no specific drug officially approved. Antibodies, particularly single-domain antibodies, represent safe and effective candidates for specific drugs against BoNT. In this study, the receptor-binding domain of botulinum toxin (BoNT/AHC_C) was utilized to immunize Bactrian camels, resulting in the generation of a nanobody phage library. From this library, a high-affinity binding antibody, designated A1, and a neutralizing antibody, named HM, were successfully obtained through SPR-based screening. The affinity constant of HM for botulinum toxin is 1.08E-11 M. Results from computer simulations indicate that HM binds at the same site as SV2C. Furthermore, experimental findings demonstrate that HM exhibits significant blocking activity at both the <i>in vitro</i> binding level and the cellular level. In mouse toxicity experiments, HM has been shown to offer protection against a 20 LD₅₀ dose of BoNT/A. Consequently, HM mitigates botulinum toxin poisoning in mice by obstructing the binding of AHC_C to SV2C.Heliyone42616https://www.ebi.ac.uk/biostudies/studies/S-EPMC11891721biostudies-literatureA novel single-domain antibody obtained from immune Bactrian camels against botulinum toxin type A using SPR-based screening method.PMC11891721Qiao CPeng SXing CChen GYu JWang JJiang ZLiu YLi XPeng FFeng JLuo LXiao HHu NLiu CWang ZfalseA novel single-domain antibody obtained from immune Bactrian camels against botulinum toxin type A using SPR-based screening method.Botulinum neurotoxin (BoNT) is a highly lethal toxin produced by the anaerobic bacterium Clostridium botulinum, which leads to nerve paralysis following poisoning. At present, there is no specific drug officially approved. Antibodies, particularly single-domain antibodies, represent safe and effective candidates for specific drugs against BoNT. In this study, the receptor-binding domain of botulinum toxin (BoNT/AHC_C) was utilized to immunize Bactrian camels, resulting in the generation of a nanobody phage library. From this library, a high-affinity binding antibody, designated A1, and a neutralizing antibody, named HM, were successfully obtained through SPR-based screening. The affinity constant of HM for botulinum toxin is 1.08E-11 M. Results from computer simulations indicate that HM binds at the same site as SV2C. Furthermore, experimental findings demonstrate that HM exhibits significant blocking activity at both the <i>in vitro</i> binding level and the cellular level. In mouse toxicity experiments, HM has been shown to offer protection against a 20 LD₅₀ dose of BoNT/A. Consequently, HM mitigates botulinum toxin poisoning in mice by obstructing the binding of AHC_C to SV2C.2025-01-01T00:00:00Z2025 Feb2026-06-01T20:12:42.687Z2025-04-07T07:50:15.411ZS-EPMC118917214006604710.1016/j.heliyon.2025.e42616