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Heterologous screening of hybridomas for the development of broad-specific monoclonal antibodies against deoxynivalenol and its analogues

In: World Mycotoxin Journal
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Y. Guo

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M. Sanders Laboratory of Food Analysis, Ghent University, Harelbekestraat 72, 9000 Ghent, Belgium

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A. Galvita Laboratory of Food Analysis, Ghent University, Harelbekestraat 72, 9000 Ghent, Belgium

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A. Heyerick Laboratory of Pharmaceutical Biotechnology, Ghent University, Harelbekestraat 72, 9000 Ghent, Belgium

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D. Deforce Laboratory of Pharmaceutical Biotechnology, Ghent University, Harelbekestraat 72, 9000 Ghent, Belgium

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M. Bracke Laboratory of Experimental Cancer Research, Ghent University, De Pintelaan 185, 9000 Ghent, Belgium

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S. Eremin Department of Chemical Enzymology, M.V. Lomonosov Moscow State University, Leniskie Gory 1, Moscow 119992, Russia

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S. De Saeger Laboratory of Food Analysis, Ghent University, Harelbekestraat 72, 9000 Ghent, Belgium

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Hapten heterology was introduced into the steps of hybridoma selection for the development of monoclonal antibodies (MAbs) against deoxynivalenol (DON). Firstly, a novel heterologous DON hapten was synthesised and covalently coupled to proteins (i.e. bovine serum albumin (BSA), ovalbumin and horseradish peroxidase) using the linkage of cyanuric chloride (CC). After immunisation, antisera from different DON immunogens were checked for the presence of useful antibodies. Next, both homologous and heterologous enzyme-linked immunosorbent assays were conducted to screen for hybridomas. It was found that heterologous screening could significantly reduce the proportion of false positives and appeared to be an efficient approach for selecting hybridomas of interest. This strategy resulted in two kinds of broad-selective MAbs against DON and its analogues. They were quite distinct from other reported DON-antibodies in their cross-reactivity profiles. A unique MAb 13H1 derived from DON-CC-BSA immunogen could recognise DON and its analogues in the order of HT-2 toxin ≯ 15-acetyl-DON ≯ DON ≯ nivalenol, with IC50 ranging from 1.14 to 7.69 μg/ml. Another preferable MAb 10H10 generated from DON-BSA immunogen manifested relatively similar affinity to DON, 3-acetyl-DON and 15-acetyl-DON, with IC50 values of 22, 15 and 34 ng/ml, respectively. This is the first broad-specific MAb against DON and its two acetylated forms and thus it can be used for simultaneous detection of the three mycotoxins.

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