sepharose and Hypergammaglobulinemia

We describe a technique for estimating the mass of anti-DNA antibodies by immunonephelometry of serum immunoglobulins (IgG, IgA, IgM) before and after adsorption onto DNA bound to agarose-polylysine columns. Sixteen patients with systemic lupus erythematosus and 16 age- and sex-matched controls were studied. Precision was determined for high-value (in 10 patients) and low-value (in nine controls) ranges for each of the immunoglobulins. Within-run CVs ranged from 3.0% (IgG, controls) to 11.8% (IgA, patients); between-run CVs ranged from 15.5% (IgG, patients) to 25.2% (IgM, patients). We found anti-DNA antibody concentrations (mean +/- SD) in systemic lupus erythematosus of 1.981 +/- 1.015 g/L for IgG (controls: 0.243 +/- 0.231, p less than 0.001), 0.257 +/- 0.215 g/L for IgA (controls: 0.038 +/- 0.035, p less than 0.001), and 0.282 +/- 0.234 g/L for IgM (controls: 0.191 +/- 0.165, p greater than 0.05). Sensitivity and linearity are such that fivefold dilutions of patients' serum with either a buffered albumin solution or control serum yielded values close to the expected values for IgG. Similarly diluted sera gave inordinately high values in the radiometric binding assay. Neither parametric (linear regression) nor nonparametric correlation methods (Spearman's rank and Kendall's tau) show a significant correlation between patients' data obtained by the present technique and that by a radiometric binding assay (p greater than 0.05), although combined data from patients and controls demonstrate a significant nonparametric correlation (p less than 0.005 for Spearman's and p less than 0.02 for Kendall's).

Topics: DNA-Binding Proteins; Humans; Hypergammaglobulinemia; Immunoglobulin A; Immunoglobulin G; Immunoglobulin M; Immunoglobulins; Immunosorbent Techniques; Lupus Erythematosus, Systemic; Nephelometry and Turbidimetry; Polylysine; Radiochemistry; Sepharose

A simple immunosubtraction technique and its application to a standard immunofixation procedure is described. Examples are presented where monoclonal immunoglobulin bands were successfully classified using immunosubtraction after other techniques had failed.

Topics: Antibodies, Monoclonal; Humans; Hypergammaglobulinemia; Immune Adherence Reaction; Immunoelectrophoresis; Immunoglobulin A; Immunoglobulin G; Sepharose

A method is described for preparative isoelectric focusing in agarose using low electroendosmotic agarose. Resolution comparable to that seen on analytical polyacrylamide gels is attainable as demonstrated by the isolation of bands with identical idiotypes from the serum of a patient with a monoclonal gammopathy.

Topics: Electrophoresis, Agar Gel; Humans; Hypergammaglobulinemia; Immunoglobulin Idiotypes; Isoelectric Focusing; Multiple Sclerosis; Polysaccharides; Sepharose