Protein A-bearing Staphylococcus aureus as the solid phase in an enzyme immunoassay and its application to...

Protein A-bearing Staphylococcus aureus was used as a solid-phase matrix in a sandwich-type enzyme immunoassay for urinary albumin. Heat-inactivated, formalin-fixed bacteria were coated with affinity-purified goat anti-human albumin, exposed to solutions containing standard or unknown concentrations of albumin, then challenged with an alkaline phosphatase/anti-human albumin conjugate obtained by periodate oxidation. Alkaline phosphatase activity bound to the bacteria was a function of albumin concentration from 25 to 1000 micrograms/L. This assay was applied to determinations of urinary albumin concentrations between 1.25 and 1000 mg/L. Between-run CV was 2.55 (63.9 mg/L concentration). Within-run CVs for albumin concentrations of 1.9, 38.1, and 638.0 mg/L were 3.7, 3.7, and 2.4%, respectively. Analytical recovery was 95 to 107% across the full working range of the assay. Bence Jones proteins and hemoglobin had no significant effect on the assay. Nonspecific binding of the enzyme-antibody conjugate was 1.3% (SD = 0.7%). Values agreed well with those by radial immunodiffusion.