Development of Capture ELISA Using a Biotinylated Monoclonal Antibody for Detection of Botulinum Neurotoxin Type A.
10.4167/jbv.2008.38.3.119
- Author:
Yun Jeong KIM
1
;
Na Ri SHIN
;
Jeong Hee KIM
;
So Yeon YOON
;
Gi Eun RHIE
;
Bong Su KIM
;
Hee Bok OH
Author Information
1. Center for Infectious Diseases, National Institute of Health, 5-Nokbun-dong, Eunpyung-gu, Seoul, Korea. hboh@nih.go.kr
- Publication Type:Original Article
- Keywords:
BoNT/A;
Capture ELISA;
Biotinylated monoclonal antibody
- MeSH:
Animals;
Antibodies, Monoclonal;
Clostridium botulinum;
Enzyme-Linked Immunosorbent Assay;
Humans;
Limit of Detection;
Mass Screening;
Mice;
Sensitivity and Specificity
- From:Journal of Bacteriology and Virology
2008;38(3):119-125
- CountryRepublic of Korea
- Language:Korean
-
Abstract:
A capture enzyme-linked immunosorbent assay (capture ELISA) was developed to detect Clostridium botulinum neurotoxin type A (BoNT/A) in assay buffer and human serum. The assay is based upon affinity-purified rabbit polyclonal and biotinylated monoclonal antibodies directed against the BoNT/A complex purified from C. botulinum ATCC19397. For the capture ELISA, the optimized amount (2 microgram/ml) of rabbit polyclonal antibody was immobilized on ELISA plates to detect BoNT/A (ranging from 0 to 500 ng/ml), which was recognized by 2 microgram/ml of the monoclonal antibody. From three independent repeated experiments, standard curves were linear over the range of 0~31.25 ng/ml BoNT/A and the coefficients (r(2)) ranged from 0.9951~0.9999 for all assays. The inter-variations were typically 0.50~6.93% and the specificity was confirmed by showing no cross-reactivity against BoNT/B and /E. The detection limit of capture ELISA was 0.488 ng/ml, which was close to mouse LD(50). In addition, application with BoNT/A-spiking human sera showed a possibility to detect BoNT/A with capture ELISA from the contaminated human sera. Taken together, the newly developed capture ELISA could serve as a rapid and sensitive screening tool for detecting BoNT/A simultaneously from massive specimens.
