Research paper A flow cytometry based competitive fluorescent microsphere immunoassay (CFIA) system for detecting up to six mycotoxins Arpad Czeh a, b, ⁎, Frank Mandy a , Szilvia Feher-Toth a, b , Livia Torok a , Zoltan Mike a , Balazs Koszegi a , Gyorgy Lustyik a, b a Soft Flow Hungary R&D Ltd. Pecs, Hungary b University of Pecs, Faculty of Medicine, Department of Biophysics, Pecs, Hungary article info abstract Article history: Received 11 March 2012 Received in revised form 18 July 2012 Accepted 18 July 2012 Available online 25 July 2012 Background: Exposure to multiple mycotoxins through the food chain represents a major potential health hazard to both humans and livestock. They can cause a variety of severe acute as well as chronic diseases. Eliminating mycotoxins from various grain crops is a global health priority. According to the Food and Agriculture Organization (FAO), world food production needs to double by 2050. Innovative solutions will be required to sustain toxin free grain supplies worldwide. Methods: A competitive flow cytometry based multiplexed assay with fluorescent micro- spheres has been developed. The new multiplexed method can analyze simultaneously any one or all six major mycotoxins. They include: Ochratoxin A (OTA), Aflatoxin B1 (AFB1), Fumonisin B1 (FB1), T-2 toxin (T-2), Deoxynivalenol (DON) and Zearalenone (ZEA), which are all potential human health hazards. The CFIA described here includes a simplified single extraction step for mycotoxins from specimens and a comprehensive post acquisition software module. The new assay system was developed with a FACSArray™ BD Bioanalyzer flow cytometer (BD Biosciences, Belgium). Results: The CFIA performs favourably when compared to commercial ELISA. Sensitivity range with CFIA increased between 13% and 100% with an average improvement of 50% for the six mycotoxins. Conclusions: The multiplexed assay presented here has the unique capacity to quantify up to six mycotoxins simultaneously from a single specimen extraction. CFIA's poly-mycotoxin detection sensitivity exceeds standard ELISA. CFIA may be part of a comprehensive assay system that will provide reliable and effective safeguard for agricultural commodities to be free of mycotoxins. © 2012 Elsevier B.V. All rights reserved. Keywords: Poly-mycotoxin environment Flow cytometry Competitive fluorescent-microsphere based immunoassay (CFIA) Acceptable maximum limits for mycotoxins (ML's) Certified reference material (CRM) 1. Introduction Global corn production is over 700 million metric tons per year. Half of the world's corn is produced in: US, China, India and EU (USDA, 2010). Harmful mycotoxins are released and produced by various species of fungi that frequently contam- inate grain (Table 1). The toxins are low-molecular weight secondary metabolic fungal products. Mycotoxins can cause a variety of acute and chronic diseases. They represent a major potential health hazard to both humans and livestock (Table 1). Pathological effects can range from immediate toxic response, endocrine abnormalities, impaired immunity, and carcinogenic and teratogenic effects (Liu and Wu, 2010; Reddy and Bhoola, 2010). In the agriculture commodities market, the elimination of mycotoxins from the food chain is a prime objective. In the future, flow cytometry based assays such as described below, may have a significant role safeguarding global food supplies. In the past 20 years, to enhance food safety, the European Commission, Food and Drug Administration of the USA (FDA), Journal of Immunological Methods 384 (2012) 71–80 ⁎ Corresponding author at: Soft Flow Hungary R&D Ltd. Pecs, Hungary. E-mail address: arpad.czeh@gmail.com (A. Czeh). 0022-1759/$ – see front matter © 2012 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.jim.2012.07.010 Contents lists available at SciVerse ScienceDirect Journal of Immunological Methods journal homepage: www.elsevier.com/locate/jim