期刊
TOXINS
卷 9, 期 4, 页码 -出版社
MDPI AG
DOI: 10.3390/toxins9040145
关键词
HT-2 toxin; recombinant antibodies; ELISA; noncompetitive; Fab; scFv; alkaline phosphatase; fusion protein; cereal grains
资金
- VTT
- Finnish Funding Agency for Technology and innovation (TEKES) [40305/2011]
- European Union Seventh Framework Programme under Graphene Flagship and Graduate School of Advanced Diagnostic Technologies and Applications (DIA-NET) [604391]
We developed an HT-2 toxin-specific simple ELISA format with a positive read-out. The assay is based on an anti-immune complex (IC) scFv antibody fragment, which is genetically fused with alkaline phosphatase (AP). The anti-IC antibody specifically recognizes the IC between a primary anti-HT-2 toxin Fab fragment and an HT-2 toxin molecule. In the IC ELISA format, the sample is added together with the scFv-AP antibody to the ELISA plate coated with the primary antibody. After 15 min of incubation and a washing step, the ELISA response is read. A competitive ELISA including only the primary antibody recognizes both HT-2 and T-2 toxins. The anti-IC antibody makes the assay specific for HT-2 toxin, and the IC ELISA is over 10 times more sensitive compared to the competitive assay. Three different naturally contaminated matrices: wheat, barley and oats, were used to evaluate the assay performance with real samples. The corresponding limits of detection were 0.3 ng/mL (13 mu g/kg), 0.1 ng/mL (4 mu g/kg) and 0.3 ng/mL (16 mu g/kg), respectively. The IC ELISA can be used for screening HT-2 toxin specifically and in relevant concentration ranges from all three tested grain matrices.
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