Solid-phase immunoassay methods for quantitation of IgG and viral antibodies in cerebrospinal fluid and its electrophoretic fractions

Abstract

A sensitive solid-phase enzyme immunoassay (EIA) for quantitative determinatio of the immunoglobulin G (IgG) was developed. It was used with a sensitive 4-layer radioimmunoassay (RIA) for the determination of viral antibodies in cerebrospinal fluids with low concentrations of antibodies. This non-competitive EIA comprised 3 main steps: (1) IgG to be assayed was adsorbed to protein A on polystyrene (PS) beads; (2) the solid-phase was then incubated with the enzyme-labelled antihuman IgG; and (3) the enzyme activity on the immunosorbent was finally measured by its effect on the substrate. The intensity of the colour was a function of the amount of IgG in the sample. This assay gave reliable and consistent results between concentrations of 15 and 1000 ng/ml. A good agreement was obtained between the EIA and the single radial immunodiffusion techniques but the EIA was about 1500 times more sensitive. The non-competitive 4-layer RIA for viral antibodies comprised 3 main steps: (1) the viral antibodies to be assayed reacted with viral antigen adsorbed onto PS beads; (2) the beads were then incubated with the rabbit IgG serum; and (3) the final incubation was with 125iodine-labelled sheep antirabbit IgG. The amount of bound radioactivity was a function of the amount of antibodies in the sample. The assays developed in the present study permitted the quantitation of IgG and viral antibodies from 1 mm slices after agarose gel electrophoresis even when only 1.5–2.0 μg IgG for each of the subsequent antibody determinations was used.