Development of a highly sensitive and specific enzyme-linked immunosorbent assay for detection of sudan I in food samples

Dan Han, Meng Yu, Dietmar Knopp, Reinhard Niessner, Mei Wu, Anping Deng

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81 Scopus citations

Abstract

A highly selective and sensitive indirect competitive enzyme-linked immunosorbent assay (ELISA) for Sudan I was developed. Two hapten derivatives with different lengths of carboxylic spacer at the azo-bound para-position were synthesized and coupled to carrier proteins. The hapten-bovine serum albumin (BSA) conjugates were used as immunogens, while the hapten-ovalbumin (OA) conjugates were applied as coating antigens. The antisera which were obtained from four immunized rabbits were characterized in terms of sensitivity and specificity. At optimal experimental conditions it was found that IC 50 and LOD values of seven pairs based on four antisera and two coating antigens were in the range of 0.3-2 ng/mL and 0.02-0.1 ng/mL, respectively. The most sensitive ELISA could be established with Sudan I-propionic acid-OA coating antigen and the antiserum which was obtained with the corresponding immunogen. The cross-reactivity values of the four antisera with Sudan II, III, and IV was estimated with 0.1-14.3%. No cross-reactivity was found with six edible colorants Sunset yellow, Amarant, Kermes, Indigotin, Bright blue and Lemon yellow, indicating high specificity for Sudan I. Six food samples were fortified with Sudan I and extracted by simple sample preparation. The methanolic extracts after dilution with methanol:water (5:95, v/v) were analyzed by the developed ELISA. Assay precision and accuracy was estimated by determination of three replicates. Acceptable recovery rates of 92.5-114% and intra-assay coefficients of variation of 5.9-24.8% were obtained. The data were validated by conventional HPLC method. As revealed, both methods were highly correlated (r= 0.9851, n = 7), demonstrating the applicability of the developed ELISA for Sudan I analysis in food samples.

Original languageEnglish
Pages (from-to)6424-6430
Number of pages7
JournalJournal of agricultural and food chemistry
Volume55
Issue number16
DOIs
StatePublished - 8 Aug 2007

Keywords

  • Analysis
  • ELISA
  • Food samples
  • HPLC
  • Polyclonal antisera
  • Sudan I

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