DEVELOPMENT OF ENZYME-LINKED IMMUNOSORBENT ASSAYS FOR 4-NITROPHENOL AND SUBSTITUTED 4-NITROPHENOLS

An enzyme-linked immunosorbent assay (ELISA) was developed for the determination of 4-nitrophenol and monosubstituted 4-nitrophenols such as 2-chloro-4-nitrophenol and 3-methyl-4-nitrophenol. An antiserum (Ab1812) was selected among eight antisera against three different corresponding immunogens. 4-(Nitrophenyl)acetyl OVA conjugate (C-OVA) as a coating antigen was selected among eight hapten-protein conjugates. Using an Ab1812 dilution of 1/1000, I50 values for 4-nitrophenol, 3-methyl-4-nitrophenol, 2-chloro-4-nitrophenol, and 2-amino-4-nitrophenol were 59, 64, 31, and 57 nM, respectively. There were no cross reactivities with other substituted phenols/nitrobenzenes, and 2- or 3-nitrophenol. The detection limit of this ELISA was 1000 times lower than that of a standard colorimetric method for the analysis of 4-nitrophenol. Under the optimized condition, this procedure can be used to determine the concentration of 4-nitrophenol and/or monosubstituted 4-nitrophenols at levels as low as 0.2 ppb in different water samples without extraction and cleanup.