Study On Enzyme-linked Immunosorbent Assay Of Heavy Metal Mercury, Copper, Lead And Cadmium

The contamination with trace elements, including mercury, cadmium,lead, arsenic, copper and zinc, are known to be toxic. The environmental release of these metals can threat human health. Contamination monitoring of heavy metal should be timely and regularly. Current technologies for measuring heavy metals can measure metal contaminations sensitively and reliably. However, these instrumental analyses require expensive facilities. In addition, the processes are relatively time-consuming. Comparing with the traditional methods,immunoassays have the advantages of quick, inexpensive and also highly sensitive and selective.In this study, the method of IC-ELISA for detection heavy metals(Hg, Cu, Cd and Pb) was developed. The results are showed as following:1. Artificial antigens for mercury(Hg-MPA-BSA and Hg-GSH-BSA)was synthesized using novel mercuric haptens. The techniques, including ultraviolet spectrometry, circular dichroism and inductively coupled plasma optical emission spectroscopy were utilized for characterizing the artificial antigens. Antibodies by rabbits immunized with the antigen was producted and properties of the antibodies studied by ELISA. The results show that: the antigen is safe for immunized animals; high titer antibodiesagainst Hg-MPA are obtained implying good immunogenicity of the antigen. The method of IC-ELISA for detecting mercury have been carried, the standard curve was “y=37.38x-30.85”, and the lower limit of detection for mecury was 12.30 ng/L, and the recoveries in the water samples were in the range of 76.0 %~124.0 %.2. Copper was conjugated to two carrier proteins using penicillin G sodium salt to synthesize artificial antigens for copper. The antigens were identified by ultraviolet spectrometry, circular dichroism and inductively coupled plasma optical emission spectroscopy. These results suggested that the artificial antigens for copper were synthesized successfully. The polyclonal antibodies of copper were produced by immunizing the New Zealand white rabbit with the copper antigen. An indirect competitive enzyme-linked immunosorbent assay based on the polyclonal antibodies was developed. The assay showed a detection limit of 7.29 ng/m L. The antibody showed high affinity with copper and low cross-reactivity with other metals. Recovery ratios of copper ions from spiked water were in the range of 78.0 %~122.0 %.3. For the preparation of lead antigens, lead ions were conjugated to carrier protein(KLH and BSA) utilizing the bifunctional chelator p-NH2-Bn-DTPA. The antibody was prepared by immunizing BABL/c rats, and the method of IC-ELISA was developed, the lower limite ofdetection for lead was 6.63 ng/L, and the recovery rate was in the range of 79.4 % ~ 126.0 %. Cadmium immunoconjugates was synthesized similarly. The antibody was prepared the immunizing BABL/c rats, and the method of IC-ELISA was developed, the lower limite of detection for cadmium was 14.98 ng/L, and the recovery rate was in the range of86.2 %~144.0 %.This paper described the synthesis and characterization of antigens for four heavy metals. The techniques, including ultraviolet spectrometry,circular dichroism and inductively coupled plasma optical emission spectroscopy were utilized for characterizing the artificial antigens. The results of identification showed that the artificial antigens had been synthesized successfully. Utilizing the artificial antigens, an indirect competitive enzyme-linked immunosorbent assay was developed. The indirect competitive ELISA can be used to detect the heavy metal ions in water samples.