Characterization of antibodies against benzo[a]pyrene with thermodynamic and kinetic constants

Antibodies of a polyclonal antiserum against benzo[a]pyrene were characterized by determining thermodynamic and kinetic constants of the antigen–antibody reaction. Label-free binding assays with optical detection based on reflectometric interference spectroscopy were performed to determine these constants. Different evaluation methods for kinetic measurements were compared. Also, cross-reactivity against two other polycyclic aromatic hydrocarbons, chrysene and pyrene, was checked. The affinity constant between the antibodies and benzo[a]pyrene in homogeneous phase was determined to be K=(5.3±0.3)×107 M−1 which was in the middle of the usual range of antibody affinities. The association rate constant for the reaction at the surface was determined to be (3.8±0.9)×105 M−1 s−1, the dissociation rate constant as (9.7±0.5)×10−3 s−1. Different evaluation methods applied to the kinetic measurements led to the same results. This antiserum would be suitable for the selective determination of benzo[a]pyrene in concentrated samples.