A 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.