Enzyme immunoassay for the quantitation of immunoglobulin M class antibodies to Salmonella minnesota R595 and Escherichia coli J5 lipopolysaccharides

Abstract

The level of human immunoglobulin (IgM) in plasma specific for the lipopolysaccharide of Salmonella minnesota R595 (R595 LPS) and Escherichia coli J5 (J5 LPS) was quantitated by an enzyme immunoassay (EIA) in which purified antigen is adsorbed directly onto polystyrene-acrylic copolymer cuvettes. Highly purified anti-J5 and R595 LPS specific IgM prepared by ion-exchange resin, gel filtration, and affinity resin chromatography were used as standards. The levels of specific IgM were determined in 200 plasma samples obtained from normal donors. Anti-R595 IgM levels varied from <30 μg/ml (91%), from 30 to 100 μg/ml (8.5%), and >100 μg/ml (0.5%). Anti-J5 IgM levels in 68% of the donor plasmas were ⩽5 μg/ml. The levels in 30.5% of the donor plasmas ranged from 6 to 100 μg/ml; the remaining 1.5% had >100 μg/ml anti-J5 IgM. Specific IgM levels in four lots of normal pooled plasma each consisting of about 10,000 L averaged 12.7 μg/ml and 13.3 μg/ml for R595 and J5, respectively. The assay was modified to quantitate rabbit plasma as well. For this purpose, the EIA has been performed on microtiter plates, and the core LPS was fixed onto the wells by chemical treatment with glutaraldehyde which results in higher stability and retention of the antigen in the wells. Specificity of the EIA was demonstrated by the absence of significant cross reactivity between R595 IgM and J5 LPS and between J5 IgM and R595 LPS, furthermore, we only observed partial adsorption (∼25%–33%) of the R595 and J5 IgM by Pseudomonas aeruginosa LPS, a wild type endotoxin. The described quantitative assay is useful for both scientific studies and clinical investigations.