Microsphere flow cytometry: A promising tool for on-site rapid screening of small-molecule harmful substances in food

• A multiplexed detection platform integrating microspheres, immunoassays, and flow cytometry. • Enables on-site, high-throughput screening with a portable flow cytometer (<15 kg). • Rapidly and simultaneously screens multiple small-molecule hazards in various food matrices. • Serves as a frontline screening tool complementing mass spectrometry confirmation. Food safety, jeopardized by pervasive small-molecule contaminants (e.g., mycotoxins, pesticides, and veterinary drug residues), demands rapid, multiplexed, and on-site detection methods to overcome limitations of conventional techniques like chromatography-mass spectrometry (costly, labor-intensive) and ELISA (single-target, low-throughput). Microsphere Flow Cytometry (MFC) addresses these challenges by synergizing microsphere-based immunocompetitive assays, fluorescent barcoding technology, and portable flow cytometry. This innovative platform achieves simultaneous detection of diverse analytes through encoded microspheres, delivering attogram-level sensitivity and analyzing dozens of samples per hour with minimal preprocessing. Key advancements include: (1) multiplexing capacityvia spectral encoding, enabling concurrent screening of various kinds of compounds; (2) portabilityvia compact flow cytometers (<15 kg), facilitating real-world deployment in non-laboratory settings; and (3) cost-effectiveness, balancing high-throughput advantages with affordability for routine screening. Challenges like matrix interference and cross-reactivity were mitigated through antibody engineering, sample extraction and dilution strategies, and matrix-matched calibrations. Future research directions will focus on: enhancing detection sensitivity through quantum dot labeling technology; adopting aptamers and primer molecules to achieve broader applications and reduce detection costs; introducing microfluidic technology to realize automation of testing processes; and establishing standardized workflows to enable seamless integration of rapid screening and validation analysis. As a high-efficiency, scalable food safety monitoring tool, MFC aligns with the global demand for on-site, high-throughput, and low-cost contaminant detection. Meanwhile, it should also integrate mass spectrometry detection to enable both faster food regulation and more precise food safety assurance.