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目的:建立检测A型肉毒毒素的酶联免疫吸附法。方法:利用重组A型肉毒毒素重链羧基端片段作为免疫原和B淋巴细胞杂交瘤技术,获得了56株高亲和力的抗A型肉毒毒素单克隆抗体(mAb),建立了检测A型肉毒毒素的双mAb夹心酶联免疫吸附法(ELISA)。结果:该方法理论检测下限达到了8.34 pg/mL,检测天然A型肉毒毒素制剂时其实际检测下限可达0.78 LD50/mL,与其他型别的肉毒毒素均未见交叉反应。通过测定基质中的回收率,发现基质中的相关的蛋白对检测具有较大的影响。结论:该方法具有灵敏度高,特异性好,使用方便等优点,具有较高的的实用价值。
OBJECTIVE: To establish an enzyme-linked immunosorbent assay for the detection of botulinum toxin type A. Methods: Fifty-six high-affinity monoclonal antibodies against botulinum toxin type A (mAb) were obtained by using recombinant human botulinum toxin type A carboxy terminal fragment as immunogen and B lymphocyte hybridoma. A monoclonal antibody against type A Botulinum toxin double mAb sandwich enzyme-linked immunosorbent assay (ELISA). Results: The detection limit of this method reached 8.34 pg / mL. The detection limit of natural botulinum toxin type A was 0.78 LD50 / mL. There was no cross reaction with other botulinum toxin types. By measuring the recovery rate in the matrix, it was found that the relevant protein in the matrix had a large influence on the detection. Conclusion: The method has the advantages of high sensitivity, good specificity, easy to use, etc., and has high practical value.