High throughput determination of BTEX by a one-step fluorescence polarization immunoassayстатья
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Дата последнего поиска статьи во внешних источниках: 18 июля 2013 г.
Авторы:Eremin S.A.,
Knopp D.,
Niessner R.,
Hong J.Y.,
Park S.J.,
Choi M.J.
Аннотация:For the rapid screening of BTEX (benzene, toluene, ethylbenzene, xylenes), a fluorescence polarization immunoassay (FPIA) was developed using the fluorescence polarization analyzer Abbott TDx. Several fluorescence-labelled tracers were synthesized by binding ethylenediamine fluorescein thiocarbamyl (EDF) to various substituted phenylcarboxylic acids. The binding of 27 tracers with two antisera that can be considered as class-specific for BTEX was investigated to select optimal tracer-antibody pairs. Significant differences were found in binding, titer, sensitivity, and assay kinetics. A best pair of anti-BTEX antiserum and EDF-labelled p-tolylacetic acid tracer was selected for FPIA. To simplify the method, an immunocomplex single reagent was prepared to detect BTEX by a one-step FPIA. One-step FPIA is a rapid homogeneous type of immunoassay that has only one pipetting step, does not need separation of free and bound analyte and can be performed at room temperature. The within-run coefficient of variation was ranged between 3.4% and 5.7%. Furthermore, if the measurement can be done at constant temperature, standards for every assay run are unnecessary. Cross-reactivity studies of petroleum compounds and a BTEX mixture indicated that p-xylene was most reactive with a limit of detection (LOD) of 0.22 mu g mL(-1) in 50 mu L of sample. The LOD for toluene and benzene was 2.1 and 11 mu g mL(-1) respectively. The immunocomplex single reagent has proven to be significantly more stable than the corresponding solutions of antibody and tracer.