A capillary-based amperometric flow immunoassay for 2,4,6-trichlorophenol

This paper describes the development of two different capillary-based heterogeneous competitive flow immunoassay formats (capillary flow injection immunoassay (CFIIA) and capillary sequential injection immunoassay (CSIIA)) for the determination of 2,4,6-trichlorophenol (2,4,6-TCP). The assays are based on the competition between the analyte and an analyte derivative labelled with the enzyme beta-galactosidase, for an anti-TCP antibody, followed by the injection of the mixture at equilibrium into a flow stream, where separation between the fractions bound and unbound to the antibody is performed in a glass capillary containing immobilised protein A. The antibody-tracer fraction retained inside the protein A capillary was measured by injection of 4-aminophenyl- beta- D-galactoside (4-APG), followed by amperometric detection of the enzymatically generated 4-aminophenol (4-AP), leading to a negative correlation between the signal and the analyte concentration.The two immunoassay formats were compared in terms of sensitivity and speed, giving IC(50) values of 1.41+/-0.03 and 1.64+/-0.07 micro g L(-1), detection limits of 0.2 and 0.4 micro g L(-1), and sample throughputs of 6 and 4 h(-1) for the CFIIA and CSIIA system, respectively.The influence of different interfering chlorophenolic compounds in the assay was minor, with only one exception (i.e. 2,4-dichlorophenol). In addition, different water matrices were tested (surface, tap, and rain water), showing that the matrix influence was negligible, except for rainwater, which resulted in a 30% increase in sensitivity. As a conclusion, the assay is suitable for the fast screening of TCP present at low concentration levels in water samples.