Development of a Novel Aptamer-Antibody Sandwich Chemiluminescent Biosensor and Its Application in the Detection of Aflatoxin B

In addressing the challenges posed by high costs, low accuracy, and cumbersome operations in mycotoxin detection, a novel aptamer-antibody sandwich chemiluminescent biosensor for detecting aflatoxin B1 (AFB1) was developed. The indirect competition between AFB1, aflatoxin B1-ovomucoid complete antigen (AFB1-OVA), and rabbit anti-ovomucoid (OVA) antibody results in the formation of a sandwich complex. This sandwich assay is linked to a horseradish peroxidase-labeled antibody, which catalyzes luminol chemiluminescence for the indirect detection of AFB1. The biosensor was designed to operate with high precision, low cost, and a low detection limit for AFB1, which is contingent upon experimental conditions such as pH, reagent concentration, temperature, and incubation time. The optimization of pH, aptamer concentration, competitive incubation time, competitive incubation temperature, and HRP-labeled antibody concentration was instrumental in achieving these objectives. Experimental findings demonstrated that the sensor's detection limit was 0.067 ng/mL, exhibiting excellent linearity (R2 = 0.99679) within the concentration range of 0.25-10 ng/mL. The recovery rate of spiked samples ranged from 94.4% to 108.05%. This sensor boasts a low detection limit, straightforward operation, and minimal cost, thus offering a novel solution for developing cost-effective, high-precision mycotoxin detection methods.