Influence of coating ligands on enzyme-linked immunosorbent assay of toluene

The specificity of conjugation site for coating ligands was investigated us ing toluene-bovine serum albumin (BSA) conjugates in which BSA was conjugat ed at the position of o-, m-, and rho -toluic acid. Toluene-BSA conjugated at rho -position showed a binding activity of about 89-95%, whereas, those conjugated at o- and m-position of toluene exhibited a binding activity of 5 and 11%, respectively. On the basis of the above result, coating ligands with various proteins (OVA, BSA, KLH) were compared by conjugating with p-t oluic acid, and toluene-KLH was considered as the best coating ligand for t his ELISA. Indirect competitive ELISA method was developed using anti-tolue ne antibody and rho -position conjugated toluene-KLH. The dose-response cur ve constructed after kinetic and optimization studies showed a 1 x 10(-4)-1 x 10(2) mM detectable response range with 0.1 muM detectability. In specif icity test of the antibody, the binding capabilities of aromatic compounds substituted with nitro-, alkyl-, chloro-, and hydroxyl group were larger ra ther than those of aromatic compounds (benzene, toluene and xylene) themsel ves. Also, tests with soil and water samples that had been spiked with tolu ene resulted in 102.7-113.7% recovery.