Flow injection chemiluminescence immunoassay based on resin beads, enzymatic amplification and a novel monoclonal antibody for determination of Hg²⁺

Abstract

In the present work, a simple and sensitive flow injection chemiluminescent competitive immunoassay was developed for the determination of mercury(II) ion (Hg(2+)) based on carboxylic resin beads, a novel specific monoclonal antibody (McAb) and HRP enzyme-amplification. Resin beads with carboxyl groups were creatively employed as supports for immobilizing more coating antigen through acylamide bonds. With a competitive-type assay mode, the Hg(2+) in solution competed with the immobilized coating antigen for the limited McAb. Then, the second antibody labeled with HRP was introduced, and an effectively increased CL was obtained, which was ascribed to the catalytic activity of HRP for the luminol-PIP-H2O2 reaction. With increasing concentration of Hg(2+), the CL of this system decreases because less HRP is present in the CL reaction. At optimal conditions, the CL signal displayed a good linear relation toward Hg(2+) in the range of 0.05-200 ng mL(-1) with a detection limit (3σ) of 0.015 ng mL(-1). The immunosensor possessed high specificity, acceptable accuracy and reproducibility, and was examined in real samples with favorable results. This immunoassay will have intriguing application prospects for the determination of other heavy metal ions and environmental contaminants.

Recent advances in flow injection analysis

“…For instance, in the system developed for chemiluminescence immunoassay of Hg(II), carboxylic resin beads, packed in a glass tube, supported immobilization of antigen to improve the sensitivity of competitive immunoassay. 267 The chemiluminescence signal was measured directly from the sorbent bed. The chemiluminescence immunoassay for sensitive detection of human prealbumin was reported with the use of nanobodies, derived from camelid heavy-chain antibodies, and immobilized onto the surface of magnetic particles.…”

Chromatographic Immunoassays: Strategies and Recent Developments in The Analysis of Drugs and Biological Agents

“…In this last example, results were obtained within 10 min, and the assay gave good agreement with an HPLC reference method [66]. A variation of the simultaneous injection method was reported for the analysis of Hg 2+ , in which an immobilized analog of the target (i.e., a protein-Hg 2+ conjugate) was combined with both samples and antibodies that could bind to this metal ion [67].…”

Flow-Injection Methods in Water Analysis—Recent Developments

“…A sampling rate of 8 h −1 was reported for very sensitive determinations with an LOD of 50 ng L −1 . Chemiluminescence detection, based on the principle of the permanganate method, was used for the determination of chemical oxygen demand in an FIA system [97], as well as for the very sensitive determination of Hg(II) in surface-and tap waters [98]. In the latter example, an immunoassay based on the use of resin beads, enzymatic amplification, and a novel monoclonal antibody with strong affinity recognition and high specificity for Hg(II), were used.…”