Screening of single-stranded DNA (ssDNA) aptamers against a zearalenone monoclonal antibody and development of a ssDNA-based enzyme-linked oligonucleotide assay for determination of zearalenone in corn

J Agric Food Chem. 2015 Jan 14;63(1):136-41. doi: 10.1021/jf503733g.


A hypotoxic immunosorbent assay for the detection of zearalenone (ZEN) was developed, by identifying a single-stranded DNA (ssDNA) aptamer with high specificity and affinity for a ZEN monoclonal antibody (mAb-ZEN). ssDNA aptamers, which could mimic ZEN epitopes, were identified using the modified systematic evolution of ligands by an exponential enrichment (SELEX) technique. The purified mAb-ZEN was coated on microtiter plates as a target recognized by the random oligonucleotide ssDNA library. The binding affinity between the aptamers and mAb-ZEN during each round was measured by the biotin–streptavidin–horseradish peroxidase system. During 15 rounds of screening, an increasing binding affinity was observed. The enriched ssDNA library binding to mAb-ZEN with high affinity was cloned, sequenced, and analyzed. One aptamer (number 46), which displays the highest affinity and specificity for the mAb-ZEN, was used to establish an indirect competition enzyme-linked oligonucleotide assay (ELONA) to measure the ZEN concentration in corn. Under optimal conditions, the regression equation for quantification of ZEN was y = −0.0778x + 0.713 (R2 = 0.9981). The detection limit and IC50 were 0.01 and 0.2 ng/mL, respectively, with a working range of 0.03–2.5 ng/mL. The recovery rates of the spiked samples in the ELONA ranged from 95 to 105%. Aptamers, which can mimic many types of low-weight analytes in agricultural products, could serve as surrogates for the development of hypotoxic, environmentally friendly immunological detection methods.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antibodies, Monoclonal* / genetics
  • Antibodies, Monoclonal* / immunology
  • Antibody Specificity
  • Aptamers, Nucleotide / chemistry
  • Base Sequence
  • DNA, Single-Stranded* / chemistry
  • Immunoenzyme Techniques / methods*
  • Molecular Sequence Data
  • SELEX Aptamer Technique / methods
  • Seeds / chemistry
  • Sensitivity and Specificity
  • Zea mays / chemistry*
  • Zea mays / genetics
  • Zearalenone / analysis*
  • Zearalenone / immunology*


  • Antibodies, Monoclonal
  • Aptamers, Nucleotide
  • DNA, Single-Stranded
  • Zearalenone