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Enhanced non-toxic immunodetection of Alternaria mycotoxin tenuazonic acid based on ferritin-displayed anti-idiotypic nanobody-nanoluciferase multimers

  • Wang, Feng
  • Li, Zhen-Feng
  • Wan, De-Bin
  • Vasylieva, Natalia
  • Shen, Yu-Dong
  • Xu, Zhen-Lin
  • Yang, Jin-Yi
  • Gettemans, Jan
  • Wang, Hong
  • Hammock, Bruce D.
  • Sun, Yuan-Ming
Publication Date
Jan 01, 2021
DOI: 10.1021/acs.jafc.1c01128
Ghent University Institutional Archive
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The non-toxic immunoassay for mycotoxins is being paid more attention due to its advantages of higher safety and cost savings by using anti-idiotype antibodies to substitute toxins. In this study, with tenuazonic acid (TeA), a kind of highly toxic Alternaria mycotoxin as the target, an enhanced non-toxic immunoassay was developed based on the ferritin-displayed anti-idiotypic nanobody-nanoluciferase multimers. First, three specific beta-type anti-idiotype nanobodies (AId-Nbs) bearing the internal image of TeA mycotoxin were selected from an immune phage display library. Then, the AId-Nb 2D with the best performance was exploited to generate a nanoluciferase (Nluc)-functionalized fusion monomer, by which a one-step non-toxic immunodetection format for TeA was established and proven to be effective. To further improve the affinity of the monomer, a ferritin display strategy was used to prepare 2D-Nluc fusion multimers. Finally, an enhanced bioluminescent enzyme immunoassay (BLEIA) was established in which the half maximal inhibitory concentration (IC50) for TeA was 6.5 ng/mL with a 10.5-fold improvement of the 2D-based enzyme-linked immunosorbent assay (ELISA). The proposed assay exhibited high selectivities and good recoveries of 80.0-95.2%. The generated AId-Nb and ferritin-displayed AId-Nb-Nluc multimers were successfully extended to the application of TeA in food samples. This study brings a new strategy for production of multivalent AId-Nbs and non-toxic immunoassays for trace toxic contaminants.

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