FOOD SCIENCE ›› 2019, Vol. 40 ›› Issue (8): 256-261.doi: 10.7506/spkx1002-6630-20171128-337

• Safety Detection • Previous Articles     Next Articles

Phage Displayed Nanobody Mediated Green Immuno-PCR for Detection of Deoxynivalenol

JIANG Dongjian, LUO Xiu’er, HE Qinghua   

  1. 1. State Key Laboratory of Food Science and Technology, Sino German Joint Research Institute, Nanchang University, Nanchang 330047, China; 2. School of Food Science & Technology, Nanchang University, Nanchang 330031, China
  • Online:2019-04-25 Published:2019-05-05

Abstract: Objective: In order to develop a highly sensitive and green immunoassay for deoxynivalenol (DON) anti-idiotypic nanobody, obtained in our previous study, was used as an alternative to enzyme-labeled antigen in a fluorescence real-time immuno-PCR system. Methods: The phage displayed nanobody (P-28), which specifically binds with anti-DON antibody, was used as a competitive antigen and the DNA encoding P-28 Nanobody was used as a target for primer design. Important experimental parameters including annealing temperature, anti-DON antibody concentration and P-28 amount were optimized. Finally, we proposed a fluorescence real-time immuno-PCR method based on indirect competition for the detection of DON. Results: The linear range of the immuno-fluorescence PCR method was 0.1–1 000 ng/mL with an IC50 of (3.96 ± 2.21) ng/mL, and the limit of detection (LOD) was 0.048 ng/mL. This method showed no crossreaction with other mycotoxins. Conclusion: This method avoids the defects of environmental pollution and operational toxicity caused by using traditional synthetic enzyme-labelled antigen and has good specificity and sensitivity.

Key words: deoxynivalenol, anti-idiotypic antibody, phage displayed nanobody, enzyme-linked immunosorbent assay, immuno-PCR

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