Development of a Microarray Lateral Flow Strip Test Using a Luminescent Organic Compound for Multiplex Detection of Five Mycotoxins

While lateral flow immunoassay (LFIA) is a simple technique that offers a rapid, robust, user friendly, and point-of-care test, its capacity for multiplex detection is rather limited. This study therefore combined the multiplexity of microarray technique and the simple and rapid characteristics of LFIA to enable simultaneous and quantitative detection of five mycotoxins, namely aflatoxin B1 (AFB1), deoxynivalenol (DON), fumonisin B1 (FUMB1), T-2 toxin (T-2), and zearalenone (ZON). In addition, we have synthesized a novel extra-large Stokes shift and strong fluorescence organic compound to be used as a reporter molecule which can be detected under UV light without light filter requirement. Many parameters including microarray spotting buffer, blocking buffer, and concentrations of mycotoxin antibodies were optimized for the microarray LFIA (μLFIA) construction. With the optimal conditions, the μLFIA could accurately and quantitatively detect multiple mycotoxins at the same time. The limits of detection of AFB1, DON, FUMB1, T-2, and ZON were 1.3, 0.5, 0.4, 0.4, and 0.9 ppb, respectively. The recoveries of these five mycotoxins were 70.7%-119.5% and 80.4%-124.8% for intra-assay and inter-assay, respectively. Combining the advantages of the novel reporter molecule and the multiplex capability of μLFIA test, this system could simultaneously detect multiple mycotoxins in one sample with high specificity and high sensitivity. Moreover, this system presents a promising affordable point-of-care platform to detect other analytes as well.