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Isoelectric focusing of monoclonal immunoglobulin G, A and M followed by detection with the avidin‐biotin system
Author(s) -
Chiodi Francesca,
Sidén Åke,
Ösby Eva
Publication year - 1985
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150060305
Subject(s) - avidin , isoelectric focusing , nitrocellulose , biotin , chemistry , isoelectric point , peroxidase , agarose , chromatography , collodion , microbiology and biotechnology , biochemistry , membrane , enzyme , biology
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.