Immunoassay by graphite furnace atomic absorption spectrometry using a metal chelate as a label

Abstract

A new immunoassay method was developed by using graphite furnace atomic absorption spectrometry and EDTA–Cd 2+ chelate as a label. The new approach combined the use of EDTA–Cd 2+ chelate labeled streptavidin (SA), biotinylated antibody or antigen, and graphite furnace atomic absorption detection for both sandwich-type immunoassay and competitive-type immunoassay. The measurements of α-fetoprotein (AFP) in human sera and bensulfuron-methyl (BSM) in water were used as the models to evaluate the usefulness of the method. The assays were carried out with 96-well microtiter plate as solid-phase carrier. After the immune reaction and biotin–streptavidin binding reaction, a solution of 0.2 M HNO 3 was added to each well to dissociate Cd 2+ in the immune complex on the solid-phase into the solution. The antigen concentration, was thus, determined by measuring the Cd 2+ absorbance of the solution with graphite furnace atomic absorption spectrometry. The present method gives detection limits of 0.12 ng ml −1 for AFP and 0.95 ng ml −1 for BSM, respectively. The coefficient variations (CVs) of the method are less than 8.0%, and the recoveries are in the range of 90–110%. The concentrations of AFP in 23 human serum samples were determined, and the results were compared with those of the independently determined by time-resolved fluoroimmunoassay method. A good correlation was obtained with a correlation coefficient of 0.993.