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Toxins 2018, 10(2), 75; doi:10.3390/toxins10020075

Preparation of Monoclonal Antibody for Brevetoxin 1 and Development of Ic-ELISA and Colloidal Gold Strip to Detect Brevetoxin 1

Key Laboratory of Pathogenic Fungi and Mycotoxins of Fujian Province, Key Laboratory of Biopesticide and Chemical Biology of Education Ministry, and School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002, China
These authors contribute equally to the work.
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Author to whom correspondence should be addressed.
Received: 26 December 2017 / Revised: 28 January 2018 / Accepted: 1 February 2018 / Published: 8 February 2018
(This article belongs to the Section Marine and Freshwater Toxins)
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Abstract

Brevetoxin-1 (BTX-1), a marine toxin mostly produced by the dinoflagellatae Karenia brevis, has caused the death of marine organisms and has had numerous toxicological effects on human health. Hence, it is very necessary to develop a rapid, economical, and reliable immunoassay method for BTX-1 detection. In this study, two kinds of complete antigen were synthesized using the succinic anhydride and isobutyl chloroformate two-step methods. Conjugate BTX-1-OVA was used as an antigen for mice immunization, and BTX-1-BSA for measuring the titer of the produced antibodies. A hybridoma cell line 6C6 stably secreting monoclonal antibody (mAb) against BTX-1 was obtained by fusing SP2/0 myeloma cells with the spleen cells from the immunized mouse. The hybridoma 6C6 was injected into the abdomen of BALB/c mice to obtain ascites, and the anti-BTX-1 mAb was harvested from ascites by precipitation with caprylic acid/ammonium sulfate (CA-AS). The anti-BTX-1 mAb was identified as an IgG1 subtype, and the cross-reactivity results showed that anti-BTX-1 mAb was highly specific to BTX-1 with the affinity of 1.06 × 108 L/mol. The indirect competitive ELISA results indicated that the linear range for BTX-1 detection was 14–263 ng/mL with IC50 of 60 ng/mL, and a detection limit of 14 ng/mL. The average recovery rate from the spiked samples was 88 ± 2% in intra-assay and 89 ± 2% in inter-assay. The limit of detection (LOD) using the colloidal gold strip was 200 ng/mL with high specificity. Therefore, the anti-BTX-1 mAb can be used to detect BTX-1 in shellfish and other related samples. View Full-Text
Keywords: Brevetoxin-1; monoclonal antibody; ELISA; colloidal gold strip Brevetoxin-1; monoclonal antibody; ELISA; colloidal gold strip
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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MDPI and ACS Style

Ling, S.; Xiao, S.; Xie, C.; Wang, R.; Zeng, L.; Wang, K.; Zhang, D.; Li, X.; Wang, S. Preparation of Monoclonal Antibody for Brevetoxin 1 and Development of Ic-ELISA and Colloidal Gold Strip to Detect Brevetoxin 1. Toxins 2018, 10, 75.

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