Influence of antibody valency in a displacement immunoassay for the quantitation of 2,4-dichlorophenoxyacetic acid.

The influence of antibody valency in a displacement immunoassay was investigated by comparing the whole antibody molecule with the corresponding Fab-fragment. The displacement immunoassay for the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) takes advantage of the cross-reactivity of monoclonal anti-2,4-D antibodies and the Fab-fragments toward immobilized 2-methyl-4-chlorophenoxyacetic acid (MCPA). Due to the low affinity of the antibodies toward MCPA (cross-reactivity of approximately 30%), the addition of 2,4-D resulted in a displacement of the antibodies or the fragments. The detection limits obtained with whole anti-2,4-D antibodies and Fab-fragments were 0.1 μg/l and 0.01 μg/l 2,4-D, respectively. The whole antibodies and the Fab-fragments show similarities, such as the cross-reactivity toward MCPA (26% and 33%), and some characteristics of the calibration curve, for example the large detection range and the sensitivity. In contrast to the bivalent antibodies, however, increasing the hapten/protein ratios of the immobilized MCPA-BSA conjugates did not affect the detection limit when using the Fab-fragments. Moreover, kinetic experiments reveal a faster displacement reaction with the Fab-fragments. A disadvantage of using the Fab-fragments is the generation of lower absorbance values in the ELISA.