Search

Monoclonal antibody-based techniques have become a useful analytical technology in the agro-food sector. Nowadays, residues of the recently registered fungicide fluopyram are increasingly being found in quality control programs. In the present study, novel chemical derivatives of this pesticide were prepared and specific and high-affinity monoclonal antibodies to fluopyram were raised for the first time. Moreover, immunoassays to fluopyram were developed in two alternative enzyme-linked immunosorbent assay formats, using homologous and heterologous assay conjugates, with limits of detection below 0.05 µg L -1 . The optimized immunoassays were applied to the analysis of fluopyram in fortified plums and grapes of four different varieties as well as in in-house prepared musts and wines. Recoveries were between 76.3% and 109.6% and coefficients of variation were below 20%. Quantification limits were well below the maximum residue limits. Immunoassay performance was statistically validated with a reference chromatographic technique using samples from fluopyram-treated plum and grape cultivars., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Immunoassays are nowadays being employed for rapid contaminant analysis in clinical, environmental, and agrochemical samples. A thorough characterization of the antibody‒antigen interaction can bring light to the immunoreagent selection process in order to develop sensitive and robust tests. Thus, determination of equilibrium and reaction rate constants is usually recommendable. However, this can be quite tricky for low molecular weight compounds, and competitive strategies are commonly followed to estimate apparent affinity values. In the present study, a collection of monoclonal antibodies to penthiopyrad was raised for the first time, and apparent equilibrium constants were assessed by the Langmuir model using three different competitive enzyme-linked immunosorbent assay formats. The obtained K D values from antibody-coated assays were quite close to the corresponding K D values calculated from surface plasmon resonance (SPR) evaluation. These studies were employed to select a pair of immunoreagents for immunoassay development. The K D value for penthiopyrad of the selected antibody obtained by SPR was 0.28 nM. The optimized direct assay showed an IC 50 value for penthiopyrad of 0.42 nM (0.15 ng mL−1) in buffer. The limit of quantification for grape, must, and wine samples was 10 ng mL−1. An excellent correlation was found when immunochemical results were compared with those from LC-MS/MS. As an application case, it was determined that 58% of penthiopyrad was still found in wine after fermentation. Image 1 • The first monoclonal antibodies to penthiopyrad are herein reported. • Antibody-antigen reaction rate constants were determined by SPR. • Thermodynamic constants were estimated by different formats of competitive ELISA. • ELISA IC 50 values approach K D values under limiting immunoreagent concentrations. • A direct immunoassay was validated to measure penthiopyrad in grapes and wine. [ABSTRACT FROM AUTHOR]