Enzyme flow immunoassay using a Protein G column for the screening of triazine herbicides in surface and waste water

A method for screening of triazine herbicides in surface and waste water is presented. The method is based on an enzyme flow immunoassay (EFIA) for the detection of the free fraction of a horse radish peroxidase (HRP)-labelled antigen (tracer). This was accomplished by trapping the bound tracer fraction in a Protein G column, allowing the residual free tracer fraction to pass and be detected spectrophotometrically after incubation with an enzyme substrate. As compared with detecting the bound tracer fraction this reduces the regeneration requirements of the Protein G column used for capturing the bound fraction and, therefore, reduces assay time. A polyclonal antibody directed against simazine showed no reactivity towards tracers that were thiopropionic acid derivatives of atrazine, simazine and terbutylazine. It had good sensitivity towards tracers using derivatives of 2-chloro-4,6-(alkylamino)-s-triazines such as atrazine and simazine. The highest sensitivity was obtained with an Et/Cl/N-C5-HRP tracer because this tracer could be used in combination with the lowest concentration of antibody. The detection limit was 0.1 μgl-1 with a linear range between 0.1 and 10 μgl-1 and an assay throughput of 12 h-1. Natural water samples from various locations in Russia were analysed for triazines and the results were compared with a previously developed fluorescein flow immunoassay for triazines. The results were further verified by supported liquid membrane (SLM) extraction combined with HPLC. The results show that the two immunoassays behave differently and that the sample matrix influences their performance, however, no false negative results were obtained. The possible reasons for the different results between the two immunoassays are discussed. (C) 2001 Elsevier Science B.V.