Multiplex chemiluminescent immunoassay for screening of mycotoxins using photonic crystal microsphere suspension array

Abstract

A novel multiplex chemiluminescent mycotoxin immunoassay suspension array system was developed by combining the silica photonic crystal microspheres (SPCMs) encoding technique and a chemiluminescent immunoassay (CLIA) method. The SPCMs were used as a carrier of the suspension array and encoded by their reflectance peak positions, which overcome fluorescence photobleaching, and the potential interference between the encoding fluorescence and detection fluorescence. Aflatoxin B1 (AFB1), fumonisin B1 (FB1) and ochratoxin A (OTA) artificial antigens were immobilized on the surfaces of SPCMs by using 3-glycidoxypropyltrimethoxysilane as a linker. Horseradish peroxidase (HRP) was used as a labeling enzyme for the secondary antibody in the enzyme-catalyze H₂O₂–luminol chemiluminescence system. The CLIA detection system was easily integrated with a multifunctional microplate reader and displayed a two to three orders of magnitude dynamic linear detection range from 0.001 to 1, 0.001 to 1, and 0.01 to 1 ng mL⁻¹ for AFB1, FB1 and OTA with 50% inhibitory concentrations (IC₅₀) of 0.01, 0.036, and 0.04 ng mL⁻¹, respectively. The recovery rates are in the range of 63.5 to 121.6% for the three mycotoxins in three kinds of spiked cereal samples. The results of detection in 12 naturally contaminated cereal samples were consistent with that of the classic enzyme-linked immunosorbent assay (ELISA) method. This proposed system is simple, rapid, low cost and high throughput for multiplex mycotoxin assay.