Development of Direct Competitive ELISA Kit for the Detection of Tetrodotoxin Using HRP Labeled Antigen
The direct competitive enzyme-linked immunosorbent assay (dcELISA) kit was developed for detecting tetrodotoxin (TTX). The working conditions of the dcELISA kit including the anti-TTX mAb coating concentration, coating method, enzyme-labeled antigen concentration, the antigen diluents, reaction time and temperature were all optimized. The result showed that mAb coating concentration was 3.72 μg/ml, it was coated at the condition of minimal power treatment of microwave oven for 3 min. The enzyme-labeled antigen concentration was 4.08 μg/ml. The competitive reaction was under the condition of room temperature 25 °C for 30 min. The half maximal inhibitory concentration (IC50) of the standard curve was 20.4 ng/ml, detection limit was 1.1 ng/ml, linear range 3.3～137 ng/ml, the intra-assay CV and inter-assay CV were 6.25% and 7.34% respectively. And recovery rate of TTX ranged from 65.0% to 93.2% with the CV of 9.41～12.77%. This method is convenient, sensitive and time-saving, hope this dcELISA kit can bring benefits and reference for TTX detection.
Zhong Cao, Yinghe He, Lixian Sun and Xueqiang Cao
Q. P. Zhong et al., "Development of Direct Competitive ELISA Kit for the Detection of Tetrodotoxin Using HRP Labeled Antigen", Advanced Materials Research, Vols. 236-238, pp. 2820-2824, 2011