Competitive and noncompetitive immunoassays for the detection of benzothiostrobin using magnetic nanoparticles and fluorescein isothiocyanate-labeled peptides
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Phage-displayed peptides have been proven to be powerful reagents for competitive and noncompetitive immunoassays. However, they are unconventional reagents, which greatly limit their analytical commercial applications and require additional reagents for detection. In this work, the peptides that specifically bind with anti-benzothiostrobin monoclonal antibody (mAb) or benzothiostrobin-mAb immunocomplex were synthesized and conjugated with fluorescein isothiocyanate (FITC) as substitutes of the phage-displayed peptides to avoid their shortcomings and extend their applications. Competitive and noncompetitive fluorescence immunoassays (FIAs) for benzothiostrobin were developed by mAb coupling with magnetic nanoparticles as concentration elements and peptides conjugated with FITC as tracers. Compared with enzyme-linked immunosorbent assays, the FIAs reduced the number of steps from 6 to 2 and analysis time from more than 5 to 1.2 h. The competitive FIA showed the half-maximal inhibition concentration (IC50) of 16.8 ng mL−1 and detection range (IC10–IC90) of 1.0–759.9 ng mL−1, while the concentration of analyte producing 50% saturation of the signal (SC50) and detection range (SC10–SC90) of noncompetitive FIA were 93.4 and 5.9–788.2 ng mL−1, respectively. The average spiked recoveries were 68.33–98.50% and 73.33–96.67% for competitive and noncompetitive FIAs, respectively. The FIAs showed good correlation with high-performance liquid chromatography for the detection of benzothiostrobin in authentic samples.
KeywordsPesticide residue Immunoassay Peptidomimetic Immunocomplex Benzothiostrobin
This work was funded by the National Key Research and Development Program of China (2017YFF0210200), the Fundamental Research Funds for the Central Universities (KYZ201618), and the National Institute of Environmental Health Sciences, Superfund Research Program (P42 ES04699). C.S.B. was funded by the United States Department of Agriculture, Agricultural Research Service, National Program project NP108, CRIS 5325-42000-049-00D, and by an interagency agreement IAA # 60-2030-5-004 with the Department of Homeland Security.
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Conflict of interest
The authors declare that they have no conflict of interest.
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