Development of monoclonal antibody-based immunoassays for the analysis of bisphenol A in canned vegetables

Instituto Interuniversitario de Investigación en Biongeniería y Tecnología Orientada al Ser Humano, Universidad Politécnica de Valencia, Spain.
Journal of Environmental Science and Health Part B Pesticides Food Contaminants and Agricultural Wastes (Impact Factor: 1.2). 08/2011; 46(6):509-17. DOI: 10.1080/03601234.2011.583871
Source: PubMed


The aim of this work was the development of monoclonal antibodies (MAbs) and highly sensitive immunoassays (ELISAs) to bisphenol A (BPA), a well-known endocrine disruptor able to migrate from the internal coating of cans to food contained inside, particularly vegetables. To produce MAbs to BPA, four synthetic compounds were conjugated to proteins and used as immunizing haptens in mice. By applying hybridoma technology, several MAbs were produced and selected. These antibodies were characterized in the conjugate-coated and in the antibody-coated formats, using both homologous and heterologous conjugates. Three indirect ELISA based on the MAbs showing the highest affinity to BPA were selected. The limit of detection of the most sensitive ELISA was 0.22 nM (0.05 ng/mL), with an I₅₀ value of around 1 nM (0.23 ng/mL). An homologous ELISA based on the MAb BPAB-11 was applied to the simple, direct determination of BPA in the liquid portion of canned artichoke, peas, and sweet corn. Only sample dilution in an appropriate saline buffer was required to minimize matrix effects and to enter the ELISA working range. Recovery and precision of the method were evaluated by spiking the liquid portion of these cans with BPA at 20, 50, and 100 ng/mL. Coefficients of variation were below 20% in most cases. With regard to recovery, the analytical data obtained were also acceptable. This immunoassay has therefore proved its potential as a new tool for the rapid, sensitive and accurate determination of BPA in canned food.

Download full-text


Available from: Juan Manclus
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: A highly specific and sensitive indirect competitive enzyme-linked immunosorbent assay (icELISA)was developed for the first time for the detection of chrysoidine, a dye banned in soybean milk film. Two haptens with different spacer arms were synthesized to produce antibodies. Both homologous and heterologous immunoassay formats were compared to enhance the icELISA sensitivity. The heterologous icELISA exhibited better performance, with an IC(50) (50% inhibitory concentration) of 0.33 ng/mL, a limit of detection (LOD, 10% inhibitory concentration) of 0.04 ng/mL, and a limit of quantitation (LOQ, 20%-80% inhibitory concentration) from 0.09 to 4.9 ng/mL. The developed icELISA was high sensitive and specific, and was applied to determine chrysoidine in fortified soybean milk film samples. The results were in good agreement with that obtained by high-performance liquid chromatography (HPLC) analyses.
    Full-text · Article · Dec 2011 · Molecules
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Enzyme-linked immunosorbent assays (ELISAs) are investigated in this work for the detection of bisphenol-A (BPA), a plastic monomer and a critical contaminant in food and environment. A series of polyclonal antibodies generated in vivo using BPA-butyrate-protein conjugate and BPA-valerate-protein conjugate were evaluated on direct and indirect competitive assay formats with five competing haptens (BPA-butyrate, BPA-valerate, BPA-crotonate, BPA-acetate, and BPA-2-valerate). Two indirect ELISAs and one direct ELISA exhibiting high sensitivity and specificity for BPA were developed. The 50 % inhibition of antibody binding (IC(50)) values were 0.78 ± 0.01-1.20 ± 0.26 μg L(-1), and the limits of detection as measured by the IC(20) values were 0.10 ± 0.03-0.20 ± 0.04 μg L(-1). The assays were highly specific to BPA, only displaying low cross-reactivity (3-8 % for the indirect assays and 26 % for the direct assay) for 4-cumylphenol (4-CP), at pH 7.2. The degree of cross-reaction of 4-CP was influenced by the antibody/hapten conjugate combination, assay conditions, and the assay format. The assays were optimized for the analysis of BPA in canned vegetables, bottled water and carbonated drinks. The limits of quantification for these three evaluated sample types, based on the spike and recovery data, were 0.5, 2.5, and 100 μg L(-1), respectively.
    Full-text · Article · Apr 2012 · Analytical and Bioanalytical Chemistry
  • [Show abstract] [Hide abstract]
    ABSTRACT: Immunochemical techniques are complementary tools to modern analytical requirements. These methods rely on the production of immunoreagents with adequate binding properties. In the present study, a rationally designed and functionalized derivative of pyrimethanil - a modern anilinopyrimidine fungicide - was synthesized in order to generate for the first time high-affinity and selective antibodies to this xenobiotic. A single coupling procedure - based on hapten activation using N,N'-disuccinimidyl carbonate and purification of the active ester - was followed to prepare both immunizing and assay conjugates. Polyclonal antibodies were produced and characterized by enzyme-linked immunosorbent assay (ELISA) in four alternative formats: one indirect and three direct competitive procedures. The selected immunoassay displayed a limit of detection of 0.024 μg L(-1), far lower than the official maximum residue limits and close to the sensitivity of regular instrumental assays. This ELISA was shown to be robust to buffer changes and tolerant to the presence of little amounts of methanol, ethanol and acetonitrile. Finally, the developed assay was applied to the analysis of pyrimethanil in carrot juice samples, and a limit of quantification of 0.040 mg L(-1) was determined.
    No preview · Article · Oct 2012 · The Analyst
Show more