Isoelectric focusing of monoclonal immunoglobulin G, A and M followed by detection with the avidin‐biotin system

Abstract

Abstract The sera of 49 subjects with IgG, IgA or IgM benign monoclonal gammopathy‐components were examined by agarose gel isoelectric focusing followed by Coomassie Brilliant Blue R‐250 staining as well as avidin‐biotin amplified double‐antibody peroxidase labelling after nitrocellulose blotting. IgG‐components gave 2–10 distinct bands within the isoelectric point (p I )‐range of pH 6.5–9.5, IgA‐components were focused into 10–15 bands with p I ‐values of pH 4.5–6.5 and the IgM‐components gave 1–2 fractions within the p I ‐range of pH 4.5–6.5. The avidin‐biotin peroxidase complex (ABC) technique exhibited some tendency to non‐specific labeling related to the avidin‐biotin component and depending on the relative amounts of the individual proteins. However, the method has a definite potential for the visualization of submicrogram quantities of immunoglobulins blotted to nitrocellulose membranes after isoelectric focusing.