Development of an enzyme-linked immunosorbent assay for toosendanin

Abstract

Enzyme-linked immunosorbent assays (ELISAs) were developed by using polyclonal antibody for toosendanin (TSN), a biopesticide from Melai toosendan Sieb. et Zucc. Their application in the determination of this analyte in spiked cabbage, tomato and apple samples was studied. The haptens, 28-hemisuccinyl-TSN (TSN-S) and 28-hemiglutaryl-TSN (TSN-G) were synthesized by using esterification. Immunogen and coating antigen were synthesized by using the mixed anhydride reaction and active ester protocol, respectively. Rabbits were immunized with TSN-G-BSA and TSN-S-BSA. Using the selected antibody and coating antigen, an indirect competitive ELISA for TSN was developed, which showed an IC 50 value of 1.023 μg mL −1, with a detection limit of 0.009 μg mL −1. A direct competitive ELISA using an enzyme tracer was also developed. The assay showed an IC 50 value of 0.840 μg mL −1 with a detection limit of 0.014 μg mL −1. Both assays displayed high cross-reactivity to a closely structurally related compound. Recoveries of TSN from both immunoassays of fortified samples ranged from 76.4% to 113.2% and 75.1% to 132.3%, respectively. Linear regression analysis showed good correlation between the TSN concentrations derived from ELISA and HPLC analyses, which suggested that the ELISA is a convenient supplementary analytical tool for monitoring TSN.