There is a strong need for the development of a sensitive biosensor for real-time monitoring mycotoxins contamination in real food samples. Here, we reported a multiplexed immunochromatographic assay (ICA) using functionalized magnetic-quantum dot (QD) as capture/detection difunctional immunoprobes that can simultaneously and sensitively detect aflatoxin B1 (AFB1), ochratoxin A (OTA), and fumonisin B1 (FB1) in food. The antibodies-modified multilayer magnetic-QD nanobeads (MagQBD) allowed effective capture of three target mycotoxins from complex solution and simultaneously detect them on one ICA strip through fluorescence signal reading. The detection sensitivity of MagQBD-ICA was markedly improved due to the signal amplification of the multilayer QD shell and target enrichment. With capture/detection strategy, multiplexed detection of AFB1, OTA, and FB1 was conducted, resulting in low detection limits of 0.42, 11.48, and 4.21 pg mL–1, respectively. The excellent accuracy, specificity, and stability of the method were further confirmed by testing actual samples including rice, peanut, corn and juices. Results suggested the great potential of MagQBD-ICA for practical detection of mycotoxins-contaminated food.