Increased sensitivity of an enzyme immunoassay (ELISA) for the determination of triazine herbicides by variation of tracer incubation time

Immunoassays for triazine herbicides were tested for their reaction to the variation of the tracer incubation time. By application of a sequential technique the measuring range of atrazine could be expanded to five decades and the total duration of the test could be reduced to about 30 min. In an optimized version a lower detection limit of 9 pmol/l (2 ng/l) was achieved. The detection limit of a sensitive immunoassay for terbuthylazine is also below the concentration limit demanded of the German drinking water regulation (100 ng/l) and reaches 130 pmol/l (30 ng/l). Short tracer incubation times did not lead to increased cross-reactivities in contrast to theoretical models [1, 2]. Different mechanisms, which could cause a shift of the center point of the calibration curve, are discussed, including kinetic considerations.Nomenclature ametryn 2-(ethylamino)-4-(isopropylamino)-6-(methylthio)-1,3,5-triazine - atrazine 2-(chloro)-4-(ethylamino)-6-(isopropylamino)-1,3,5-triazine - deethylatrazine 2-(amino)-4-(chloro)-6-(isopropylamino)-1,3,5-triazine - DMSO dimethylsulfoxide - DOC dissolved organic carbon - ELISA enzyme-linked immunosorbent assay - glyme 1,2-dimethoxyethane - hydroxyatrazine 2-(ethylamino)-4-(hydroxy)-6-(isopropylamino)-1,3,5-triazine - PBS phosphate buffered saline - propazine 2-(chloro)-4,6-bis(isopropylamino)-1,3,5-triazine - simazine 2-(chloro)-4,6-bis(ethylamino)-1,3,5-triazine - terbuthylazine 2-(tert-butylamino)-4-(chloro)-6-(ethylamino)-1,3,5-triazine - TLC thin-layer chromatography - TMB 3,3,5,5-tetramethylbenzidine - tracer enzyme (peroxidase) labeled hapten