Development and Comparison of Three Diagnostic Immunoassay Formats for the Detection of Azoxystrobin

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Abstract
The currently accepted method of detection for azoxystrobin, a strobilurin fungicide, involves a labor-intensive organic solvent extraction and gas chromatography analysis. Three diagnostic assay formats, i.e., enzyme-linked immunosorbent assay (ELISA), fluorescence polarization (FP), and time-resolved fluorescence (TR-FIA), were developed and compared with regard to detection and quantification of azoxystrobin in grape extract and river, lake, and well water samples. These three assay formats require no initial sample extraction and were not affected by any of the environmental matrices tested, and each had a linear working range of 0−400 pg/mL. The polyclonal antibodies used for each of the immunoassays were specific to azoxystrobin; that is, the highest cross-reactivity to other pesticides observed was 5.7%. The limits of detection of the immunoassays were similar at 3 (ELISA), 46 (FP), and 28 (TR-FIA) pg/mL, as were the respective IC50 values of 306, 252, and 244 pg/mL. Each of the three immunoassays developed was less labor-intensive and approximately 100-fold more sensitive than the gas chromatographic method. While the three formats were comparable in terms of performance, the fluorescence polarization assay was the least labor-intensive and required the least time to perform. Keywords: Azoxystrobin; ELISA; fluorescence polarization; time-resolved fluorescence; immunoassay

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