Synthesis of haptens of organophosphorus pesticides and development of enzyme-linked immunosorbent assays for parathion-methyl

Department of Biochemistry and Institute of Natural Sciences, Yeungnam University, Gyongsan 712-749, South Korea
Analytica Chimica Acta (Impact Factor: 4.52). 09/2003; 493(1):47-62. DOI: 10.1016/S0003-2670(03)00793-1

ABSTRACT A simple synthetic method for haptens of organophosphorus (OP) pesticides with a spacer arm (aminocarboxylic acid) attached at the pesticide thiophosphate group was developed. While the previous synthetic approach for this type of haptens requires seven steps, the present method involves only two steps. Using this method, four haptens of the OP insecticide parathion-methyl were synthesized. Rabbits were immunized with either one of the two haptens coupled to bovine serum albumin for production of polyclonal antibodies. Using the serum with the highest specificity, an antigen-coated ELISA was developed, which showed an IC50 of 6.4 ng/ml with a detection limit of 0.2 ng/ml. An antibody-coated ELISA using an enzyme tracer was also developed, which showed an IC50 of 3.5 ng/ml with a detection limit of 0.3 ng/ml. The antibodies showed negligible cross-reactivity with other OP pesticides tested except with the insecticides parathion and paraoxon only in the antigen-coated ELISA.

  • Source
    • "0.3 Â 10 À3 mM) of BSA were dissolved in 2 mL of borate buffer (0.2 M, pH 8.7) to which 0.4 mL of DMF was added. A solution of an active ester (0.018 mM) in 0.1 mL of DMF was then added to the stirred protein solution, and stirring was continued for one day at 4 °C (Kim et al., 2003; Liu et al., 2007). 3.6. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Agent orange herbicides, organophosphate and triazinic pesticides analysis in olive oil and industrial oil mill waste effluents using new organic phase immunosensors a b s t r a c t New immunosensors working in organic solvent mixtures (OPIEs) for the analysis of traces of different pesticides (triazinic, organophosphates and chlorurates) present in hydrophobic matrices such as olive oil were developed and tested. A Clark electrode was used as transducer and peroxidase enzyme as mar-ker. The competitive process took place in a chloroform–hexane 50% (V/V) mixture, while the subsequent enzymatic final measurement was performed in decane and using tert-butylhydroperoxide as substrate of the enzymatic reaction. A linear response of between about 10 nM and 5.0 lM was usually obtained in the presence of olive oil. Recovery tests were carried out in commercial or artisanal extra virgin olive oil. Traces of pesticides were also checked in the oily matrix, in pomace and mill wastewaters from an indus-trial oil mill. Immunosensors show good selectivity and satisfactory precision and recovery tests per-formed in olive oil gave excellent results.
    Food Chemistry 02/2015; 169:358-365. DOI:10.1016/j.foodchem.2014.07.137
  • Source
    • "Accordingly, the generated antibodies generally show low specificity and high degree of cross-reactivity with other similarly structured molecules. Design of a hapten and a suitable carrier protein are therefore critical to ensure successful generation of antibodies having the desired specificity for therapeutic monitoring and immunodiagnostic studies (Kim, 2003). Here, we report first time a novel method for synthesis of hapten using a bacterial isolate for the preparation of hapten–protein conjugate for specific antibodies generation against commonly used herbicide diuron (3-(3,4-dichlorophenyl)-1,1-dimethylurea). "
    [Show abstract] [Hide abstract]
    ABSTRACT: A Gram-positive, Micrococcus sp. strain PS-1 isolated from diuron storage site was studied for its capability of biotransformation of phenylurea herbicide diuron to a secondary metabolite, 1-(3,4-dichlorophenyl)urea (DCPU) for bioconjugation and antibody development applications. The metabolite formed associated with profound changes in bacterial cell morphology demonstrated increase in the degradation kinetics of diuron in presence of small quantity of a surfactant. The synthesized metabolite identified by chromatographic and mass spectrometry techniques was conjugated with carrier protein, and used as an immunogen for antibodies production. The generated antibody was highly specific, demonstrating excellent sensitivity against diuron. The antibody was used as receptor molecules in standard fluorescence immunoassay (FIA) format showing detection limit of 0.01 ng/mL in the optimum working concentration range of diuron with good signal precision (∼2%). The study presented first time the degradation pathway of herbicide by specific microorganism to synthesize hapten for bioconjugation and immunoassay development.
    Bioresource Technology 10/2010; 102(3):3119-25. DOI:10.1016/j.biortech.2010.10.076
  • Source
    • "Enzyme immunoassays are widely used in medicine, food, cosmetics fields to help rapid, simple, accurate and specific, but also cost-effective, quantification of many biologically essential small molecules. However, such technology needs to be improved or designed especially for rare haptens like hormones, drugs, insecticide, marine toxins or mycotoxins (Kim et al., 2003; Brun et al., 2004; Tomasz et al., 2009). The first challenge for researchers is to develop a method to prepare effective immunogens by coupling haptens to carrier proteins or poly-L-lysine Chun et al. 7057 (PLL). "
    AFRICAN JOURNAL OF BIOTECHNOLOGY 01/2009; 8(24). DOI:10.4314/ajb.v8i24.68793
Show more


Available from