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RESEARCH ARTICLE

High Throughput Determination of BTEX by a One-Step Fluorescence Polarization Immunoassay

Sergei A. Eremin A D , Dietmar Knopp B , Reinhard Niessner B , Ji Youn Hong C , Song-Ja Park C and Myung Ja Choi C
+ Author Affiliations
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A Faculty of Chemistry, Moscow State University, Leninsky Gory 1, Moscow 119992, Russia.

B Institute of Hydrochemistry and Chemical Balneology, Technical University of Munich, 81377 München, Germany.

C Bioanalysis and Biotransformation Research Centre, Korea Institute of Science and Technology, Cheongryang Seoul 130-650, Seoul, Korea.

D Corresponding author. Email: eremin@enz.chem.msu.ru

Environmental Chemistry 2(3) 227-234 https://doi.org/10.1071/EN04082
Submitted: 22 December 2004  Accepted: 11 June 2005   Published: 27 September 2005

Environmental Context. Benzene, toluene, ethylbenzene, and xylenes (BTEX) are used as solvents in paints and coatings and are constituents of petroleum products. BTEX can contaminate air, water or soil and is toxic; benzene, in particular, is a recognized human carcinogen. Most existing methods for detecting BTEX are time-consuming, complicated and very expensive for routine screening. A rapid immunoassay of BTEX is presented that greatly simplifies environmental monitoring of water contamination.

Abstract. 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 µg mL−1 in 50 µL of sample. The LOD for toluene and benzene was 2.1 and 11 µg mL−1 respectively. The immunocomplex single reagent has proven to be significantly more stable than the corresponding solutions of antibody and tracer.

Keywords. : immunoassay — BTEX — immunodetection


Acknowledgments

This project was supported by a fund of the National Research Laboratory Program (M1-9911-00-0085), MOST, The Republic of Korea, and by NATO grant EST.CLG.978618.


References


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