Use of Rotofor in two-dimensional electrophoretic analysis: identification of a 100 kDa monoclonal IgG heavy chain in myeloma serum.
After preparative nondenaturing isoelectric focusing in a Rotofor of 1 mL human serum which showed on two-dimensional electrophoresis a monoclonal kappa light chain pI < 5.0, the monoclonal kappa light chain was identified in the tubes with pH 7.29-pH 6.96, indicating that the chain was not free and was most likely part of an immunoglobulin molecule. The combined fractions of the Rotofor were concentrated back to 1 mL and aliquots were separated by two-dimensional electrophoresis, followed by Western blotting. Enzyme immunoassay of the blot with A' human IgG (gamma chain specific) revealed a monoclonal heavy chain at a molecular size of approximately 100 kDa. Diffuse areas at 50 kDa (the attributed size of gamma heavy chain), 100 kDa, and 150 kDa were stained also, but the strong, restricted (picket fence) bands indicating a monoclonal pattern were at 100 kDa. The monoclonal light chain appears to be part of a large or dimeric IgG molecule.
Goldfarb
M F
MF
Anatek-EP, Portland, ME 04103.
D000911:Antibodies, Monoclonal / Q000097:blood*; D015153:Blotting, Western; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D006801:Humans; D007124:Immunoenzyme Techniques; D007074:Immunoglobulin G / Q000097:blood*; D007143:Immunoglobulin Heavy Chains / Q000097:blood*; D007145:Immunoglobulin kappa-Chains / Q000097:blood; D007525:Isoelectric Focusing / Q000295:instrumentation / Q000379:methods*; D007526:Isoelectric Point; D008970:Molecular Weight; D009101:Multiple Myeloma / Q000276:immunology*
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007145:Immunoglobulin kappa-Chains