Development and Comparative Study of Different Immunoenzyme Techniques for Pesticides Detection

Abstract

Different ELISA techniques have been developed for the detemination of four widely used pesticides: 2,4-dichlorophenoxyacetic acid (2,4-D), 2,4,5-trichlorophenoxyacetic acid (2,4,5-T), simazine and atrazine. Dependences between the assay scheme and the limiting detectable concentration of the pesticide were studied. The cases of preferential applying of the scheme with immobilized antibodies or one with immobilized pesticide-protein conjugate have been revealed. The following approaches resulting in lowering of ELISA sensitivity were proposed: preliminary incubation of the tested sample with antibodies, immobilization of antibodies via staphylococcal protein A, usage of monovalent fragments of antibodies instead of native ones and chemical modification of the pesticide molecules in the sample. Optimal combinations of these approaches permitted to lower the detection limit of the assays in about 5–30 times. The achieved sensitivities were 3 ng/mL for 2,4-D, 5 ng/mL for 2,4,5-T, 0.05 ng/mL for simazine, and 0.1 ng/mL for atrazine, being acceptable for purposes of ecological monitoring.

Key Words:

• Pesticides
• enzyme immunoassay
• optimization
• sensitivity
• phenoxyacetic acids
• triazines