| Autor: |
Saeed AFUH; Fujian Key Laboratory of Pathogenic Fungi and Mycotoxins, Key Laboratory of Biopesticide and Chemical Biology of the Education Ministry, School of Life Science, Fujian Agriculture and Forestry University, Fuzhou 350002, China. abdullahfarhan@hotmail.com., Ling S; Fujian Key Laboratory of Pathogenic Fungi and Mycotoxins, Key Laboratory of Biopesticide and Chemical Biology of the Education Ministry, School of Life Science, Fujian Agriculture and Forestry University, Fuzhou 350002, China. Lsmpu2008@163.com., Yuan J; Fujian Key Laboratory of Pathogenic Fungi and Mycotoxins, Key Laboratory of Biopesticide and Chemical Biology of the Education Ministry, School of Life Science, Fujian Agriculture and Forestry University, Fuzhou 350002, China. yjmail2008@126.com., Wang S; Fujian Key Laboratory of Pathogenic Fungi and Mycotoxins, Key Laboratory of Biopesticide and Chemical Biology of the Education Ministry, School of Life Science, Fujian Agriculture and Forestry University, Fuzhou 350002, China. wshyyl@sina.com. |
| Abstrakt: |
Domoic acid (DA) is a potent toxin, marine biotoxin, and primarily produced by Pseudo-nitzschia . The DA hapten was coupled with bovine serum albumin (BSA), and ovalbumin (OVA) as carrier proteins. DA-BSA conjugate was used as immunogen and DA-OVA as coating antigen. Cell fusion between spleen cells and sp2/0 myeloma cells developed 1C3 hybridoma clone producing 1C3 monoclonal antibody (mAb). Hybridoma was injected into the mice to produce ascites, and further purified by caprylic acid/ammonium sulfate method. The mAb was of IgG3 subclass, and was specific to DA with high affinity (2.5 × 10⁸ L/mol). Moreover, western blot exhibited significant specificity to the DA antigens. Indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) showed DA working range of 0.006-0.2 ng/mL. The IC 50 was 0.03 ng/mL with low limit of detection (LOD) of 0.006 ng/mL. Average DA recovery from spiked shellfish extract was 100.56% ± 2.8% with the coefficient variation of 0.01-0.1%. Hence, mAb producing 1C3 hybridoma was successfully developed and could be used to detect DA in contaminated samples. |
