Abstract
An effective biotin–streptavidin amplified enzyme-linked immunosorbent assay (BA-ELISA) was optimized and characterized for the rapid detection of Ractopamine (RAC) residue in muscular tissue. Purification of the RAC antiserum by protein A-Sepharose 4B followed with bovine serum albumin (BSA)-Sepharose 4B affinity chromatography enhanced the sensitivity and reduce the background adsorption. Blocking with 0.5% skimmed milk power and diluting streptavidin–HRP conjugates with 0.5% BSA/phosphate-buffered saline (PBS) effectively remove the nonspecific adsorption in biotin–streptavidin amplified ELISA system. The established method allowed RAC determination with an IC50 value of 0.3 ± 0.02 ng ml−1 and a limit of detection of 0.02 ± 0.003 ng ml−1, more sensitive than the other reported methods. The variation coefficients of intra-assay and inter-assay were all below 7%. RAC residue in pig muscular tissue could be quantified without matrix effects after a 5-fold extraction and 2-fold dilution with PBS. Recoveries of RAC in pig muscular tissue ranged from 75% to 82.75%. The results were also compared with those from HPLC and a good correlation was obtained (r 2 = 0.9822). The characters show that the established biotin–streptavidin amplified ELISA could be potentially useful in rapid detection of RAC in animal-derived foods.
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Acknowledgements
The authors are grateful for financial support from the Minister of Science and Technology of the People’s Republic of China (Project No. 2009BADB9B03) and the Ministry of Science and Technology of Tian** (Project No. 09JCZDJC19400)
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Zhang, Y., Gao, X., Gao, A. et al. A Biotin–Streptavidin Amplified Enzyme-Linked Immunosorbent Assay with Improved Sensitivity for Rapid Detection of Ractopamine in muscular tissue. Food Anal. Methods 5, 1214–1220 (2012). https://doi.org/10.1007/s12161-012-9363-0
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DOI: https://doi.org/10.1007/s12161-012-9363-0