Simultaneous Screening of Fumonisin B1, Aflatoxin B1, and Zearalenone by Line Immunoblot: A Computer-Assisted Multianalyte Assay System

Abstract

Abstract A line immunoblot assay was developed for the simultaneous screening of fumonisin B1 (FB1), aflatoxin B1 (AFB1), and zearalenone (ZEA). Monoclonal antibodies for each of these toxins were immobilized as multiple lines on nitrocellulose membrane strips and sectored into hydrophobic compartments to minimize use of reagents. A modified enzyme-linked immunosorbent assay was conducted whereby free mycotoxins and horseradish peroxidase-labeled mycotoxins competed for binding to the nitrocellulose-bound antibodies. Color intensity of lines formed by a precipitating substrate was inversely related to mycotoxin concentration. Detection limits for the individual mycotoxins, as determined by visually comparing the color intensity of positive strips to negative controls, were 500, 0.5, and 3 ng/mL for FB1, AB1, and ZEA, respectively. Line density was quantitatively assessed also by using a camera, video monitor, and microcomputer equipped with a video-digitizing board. The assay could be used to determine range values for the various mycotoxins in extracts of spiked corn in less than 30 min, and values could be recorded on the microcomputer hard disk. This combination of line immunoblot assay and image analysis, termed computer-assisted multianalyte assay system, should be applicable to the simultaneous screening of multiple mycotoxins and other agricultural residues in food.