Mycotoxin contamination in the food chain has caused serious health issues in humans and animals. Thus, a rapid on-site and lab-independent detection method for mycotoxins, such as aflatoxins (AFTs), is desirable. Microfluidic chip based immunosensor technology is one of the most promising methods for fast mycotoxin assays. In this review, we cover the major microfluidic immunosensors used for mycotoxin analysis, via flow-through (capillary electromigration) and lateral flow technology. Sample preparation from different matrices of agricultural products and foodstuffs is summarized. The choice of materials, fabrication strategies, and detection methods for microfluidic immunosensors are further discussed in detail. The sensors application in mycotoxin determination is also outlined. Finally, future challenges and opportunities are discussed.