The Study Based On Immunoassays For Rapid Detection Of Sudan Ⅰ In Food

Two methods based on immunoassays were developed for rapid detection of Sudan I in food by preparing anti-Sudan I polyclonal antibodies. The assays were enzyme-linked immunosorbent assay and colloidal gold immunochromatographic method.Sudan I derivate was synthesized via diazotization measures, and Sudan I-gelatin immunogen was obtained by mixed acid ahhdride method to couple Sudan I derivative with gelatin. The immunoantigen was injected into new Zealand white rabbits to prepare the anti-Sudan I polyclonal antibodies. The hyperimmune serums were collected when the immune programme was finished, and purified via octanoicacid ammoniumsulfate precipitation and affinity chromatography. Finally, the specific anti-Sudan I antibodies were obtained.An indirect ELISA was applied to test the titer and affinity of Sudan I antibodies.Results showed the titer of antibody was 1: 32 000. The limit detection was 1.0 jig/L and IC50 was 15.0 jig/L by the inhibition ratio curve, which proved the antibodies have high affinity. The recoveries were among 85.88%-97.83%, and coefficients of variation were from 4.9% to 7.4%. The cross-reactivity for Sudan II, III, IV were 2.7%, 6.5% and 4.1%, respectively.During the process of immunochromatographic (IC) strip, the reaction was based on the nitrocellulose (NC) membrane, and the Sudan I antibody and anti-BSA antibody were immobilised on the NC membrane as the test reagent and control reagent, respectively. The goId-labled Sudan I antigen was coated on the glassfiber. With the help of chromatograph affect, the possible Sudan I in samples and the go Id-labeled Sudan I-BSA conjugate would competitively capture the immobilised antibody. The more Sudan I present in the samples, the more effective competition reaction with gold-labeled conjugate occurred. The limit detection was found to be 10 fig/kg in chili powder by optimizing the concentrations of anti-Sudan I antibodies and anti-BSA antibodies. Quantitative analysis of Sudan I could be achieved between 5.0 fig/kg and 20.0 jig/kg. Low cross reactivity was found among other Sudan dyes. Compared with HPLC methods, the results showed a good correlation with a square of correlation coefficient of 0.941. The analysis was high performance, concenient and could be completed in shorter time, suitable for rapid detect Sudan I residue in food and preliminary screening detections for abundant samples.