Development of ELISA for Detection of Mercury Based on Specific Monoclonal Antibodies Against Mercury-Chelate

Immunoassays for heavy metals offer an alternative approach to traditional techniques for detection of mercury. In this study, a mercury-chelate was prepared with 1-(4-aminobenzyl) ethylenediamine-N,N,N′,N′-tetraacetic acid (aminobenzyl-EDTA). The resulting complex was linked to keyhole limpet hemocyanin (KLH) or bovine serum albumin via the amino group and used as the immunizing antigen or detection antigen, respectively. BALB/c mice were immunized with KLH-aminobenzyl-EDTA-Hg and spleen cells from BALB/C mice were fused with Sp2/0 cells. One cell line (5F7) produced monoclonal antibodies with preferential selectivity and sensitivity for aminobenzyl-EDTA-Hg. This cell line had an affinity constant of 4.31 × 109 L/mol and its cross-reactivity (CR) with other metals was <2%. The antibody was used for competitive indirect ELISA (CI-ELISA) for Hg2+ measurements. The detection range was 0.087–790.4 μg/L and the lower limit of detection was 0.042 μg/L. The concentrations of mercury in environmental water samples obtained by CI-ELISA correlated well with graphite furnace atomic absorption spectrometry (GFAAS), and the mean recovery was 88.82% to 104.64%. These results indicate that this method could be used for monitoring mercury of water.