No abstract

Strobilurin fungicides are nowadays among the most important fungicides in the market of active agrochemicals. Pyraclostrobin, which belongs to the last generation of this family of molecules, shows a broader antifungal activity spectrum and higher efficiency and security profiles than previous fungicides. This paper describes the synthesis of functionalized haptens, the production of monoclonal antibodies, and the development of enzyme-linked immunosorbent assays (ELISA) for the detection of pyraclostrobin. A conformational analysis of hapten structure was performed, which provided relevant data concerning the length of the spacer arm. A very useful strategy has been followed for the screening of hybridomas, leading to the selection of a panel of high-affinity monoclonal antibodies to pyraclostrobin. Moreover, different immunoassays have been characterized using the conjugate-coated indirect ELISA format, and limits of detection below 0.1 microg/L have been obtained. Also, a simplified one-step procedure has been carried out with two indirect assays. Finally, these results have been compared with the performance of the same antibodies in the antibody-coated direct ELISA format.

The development of immunoassays for the detection of the plant growth regulator forchlorfenuron (CPPU) is described. To achieve that purpose, a set of CPPU derivatives has been obtained by the previous synthesis of the adequate p-aminophenyl alkanoic acid. Protein conjugates of these compounds have been used as immunogens to produce rabbit polyclonal antibodies and a collection of mouse monoclonal antibodies. Additionally, a battery of structural analogues of the target analyte has been synthesized and used for the characterization of antibody binding. This strategy has demonstrated that most antibodies followed Landsteiner's principle, although some monoclonal antibodies showing important deviations from this behavior have also been found. Finally, different assay formats have been developed with a variety of antibodies and conjugates, and a rapid procedure has been optimized for the indirect ELISA format using monoclonal and polyclonal antibodies. In the indirect competitive ELISA, assay IC50 values for CPPU below 0.5 nM were found with LODs as low as 0.013 nM.