[Immunonanogold catalytic spectrophotometric determination of trace IgG]

Gold nanoparticles the size of about 10 nm were prepared by improved trisodium citrate reduction procedure, and were used to label goat anti-human IgG to obtain a sensitive spectral probe for IgG in the condition of pH 6.5. The immune reaction of nanogold-labeled IgG antibody (anti-IgG) with the antigen IgG took place to form the nanogold immune complex in pH 7.O Na2HPO4-C6H8O7 buffer solution and in the presence of polyethylene (PEG). The optimal immunoreaction conditions were pH 7.0, 10.76 microg x mL(-1) nanogold-labeled anti-IgG, 8.0% PEG 6000 and incubation time of 30 min under the ultrasonic irradiation. After centrifuging for 15 min at 16000 rpm, the excess nanogold-labeled anti-IgG in the upper solutions was obtained, and was used to catalyze the colored particle reaction of HAuCl4 with NH2 OH x HCl to produce gold particles with bigger size. The influence of pH value, HAuCl4 and NH2OH x HCl concentration, and reaction temperature and time on the immunonanogold catalytic reaction was considered spectrophotometrically. A pH 2.27 Na3C6H5O7-HCl buffer solution, 0.094 mmol x L HAuCl4, 1.92 mmol x L NH22OH x HCl, and reaction time of 6 min at 30 degrees C water bath were chosen for use. Results demonstrated that with increasing IgG, the concentration of gold labeled anti-IgG in the upper solution decreased, and the absorbance decreased linearly. Linear relationships between the decreased absorbance at 700 nm and the IgG concentration CIgG in the range of 0.10-10 ng x mL(-1) were obtained. Its regress equation was deltaA(760 nm) = 0.0144c(IgG) + 0.0042, the related coefficient was 0.9926, and the detection limit reached 0.06 ng x mL(-1) IgG. The influence of foreign substances on the determination of 3 ng x mL(-1) IgG was examined, with the relative error +/-10%. Results showed that the following coexistent substances had no impact on the assay: 6000 ng x mL(-1) HSA, 6000 ng x mL(-1) gluocose, 6000 ng x ml(-1) Zn(II), 3000 ng x mL(-1) IgA, 3000 ng x mL(-1) Ca(II), 3000 ng x mL(-1) L-arginine, 3000 ng x mL(-1) beta-phenylalanine, 2400 ng x mL(-1) Cu(II), 2400 ng x mL(-1) EDTA, 2400 ng x mL(-1) L-cystinol etc. This showed that the assay has high selectivity. The sensitive, rapid and highly specific assay was applied to the quantification of IgG in human sera, with satisfactory results.