基于单克隆抗体的酶免疫分析法用于检测A型肉毒杆菌神经毒素

Monoclonal antibody-based enzyme immunoassay for detection of botulinum neurotoxin type A.

作者信息

Chiao Der-Jiang, Wey Jiunn-Jey, Tang Shiao-Shek

机构信息

Institute of Preventive Medicine, National Defense Medical Center, Taipei, Taiwan, Republic of China.

出版信息

Hybridoma (Larchmt). 2008 Feb;27(1):43-7. doi: 10.1089/hyb.2007.0529.

DOI:10.1089/hyb.2007.0529

PMID:18294076

Abstract

A sensitive and specific ELISA was developed to detect BoNT/A in biological fluids. The assay is based on the sandwich format using monoclonal antibodies (MAb) of two distinct specificities. An affinity-purified anti-BoNT/A heavy chain MAb (150-3) is utilized to adsorb BoNT/A from solution; the second anti-BoNT/A heavy chain MAb (44-1A) conjugated with peroxidase is then used to form a sandwich. Peroxidase allows color development and measurement of optical density at 450 nm. Standard curves were linear over the range of 2.5 to 100 ng/mL BoNT/A. The limit of detection was below 5 ng/mL in assay buffer, as well as in a 1:10 dilution of urine or 1:50 dilution of human serum spiked with BoNT/A. The developed BoNT/A assay also showed no cross-reaction to type B neurotoxin (BoNT/B) and type E neurotoxin (BoNT/E).

摘要

开发了一种灵敏且特异的酶联免疫吸附测定法（ELISA）来检测生物体液中的肉毒杆菌毒素A（BoNT/A）。该测定法基于夹心模式，使用两种不同特异性的单克隆抗体（MAb）。一种亲和纯化的抗BoNT/A重链单克隆抗体（150-3）用于从溶液中吸附BoNT/A；然后使用与过氧化物酶偶联的第二种抗BoNT/A重链单克隆抗体（44-1A）形成夹心结构。过氧化物酶可实现显色并测量450nm处的光密度。在2.5至100ng/mL BoNT/A范围内，标准曲线呈线性。在测定缓冲液中以及在加入BoNT/A的1:10稀释尿液或1:50稀释人血清中，检测限均低于5ng/mL。所开发的BoNT/A测定法对B型神经毒素（BoNT/B）和E型神经毒素（BoNT/E）也未显示交叉反应。