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Determination of Amaranth in Beverage by Indirect Competitive Enzyme-linked Immunosorbent Assay (ELISA) Based on Anti-amaranth Monoclonal Antibody

Zhang, Bo, Du, Daolin, Meng, Meng, Eremin, Sergei A., Rybakov, Victor B., Zhao, Junhong, Yin, Yongmei, Xi, Rimo
Food analytical methods 2014 v.7 no.7 pp. 1498-1505
antigens, azo dyes, beverages, cross reaction, detection limit, enzyme-linked immunosorbent assay, erythrosine, food coloring, inhibitory concentration 50, monitoring, monoclonal antibodies, risk, transport proteins
Amaranth (E 123) is a member of azo dyes, and it is allowed to use in various foods. The acceptable maximum addition of amaranth is strictly fixed because of its potential risk to physical health. The objective of this study was to prepare a specific anti-amaranth monoclonal antibody and develop an indirect competitive ELISA for amaranth quantification analysis. The immunogen and the coating antigen were designed by introducing a carboxyl group into amaranth for the conjugation with carrier proteins. Based on the immunogen, the monoclonal antibody exhibits satisfactory performances and the proposed ELISA shows an IC₅₀of 20.33 ng mL⁻¹. The limit of detection is as low as 3.35 ng mL⁻¹, and the linear standard curve of the method ranges from 3.0 to 243.0 ng mL⁻¹. Additionally, the antibody reflects minimal cross-reactivity (<1 %) with six related food dyes (erythrosine, ponceau 4R, allura red, tartrazine, sunset yellow FCF, and brilliant blue). The recoveries of amaranth spiked beverage samples are in the range of 85.8–100.7 % with low coefficient of variation values (<11.5 %). The data shows that the developed ELISA provides a simple, sensitive, specific, and accurate alternative for amaranth determination and monitoring. Furthermore, it is the first time that icELISA of amaranth is developed based on monoclonal antibodies.