Micropurification and two-dimensional polyacrylamide gel electrophoresis of immunoglobulins for studying the clonal diversity of antigen-specific antibodies.
Presented here is a simplified method for purifying antigen-specific antibodies and analysing them by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE). For this procedure, specific immunoglobulins (Ig) are bound to resin beads coated with antigen and, in the presence of sodium dodecyl sulphate and dithioerythritol, loaded directly onto isoelectric focusing strips for 2D-PAGE analysis. This technique bypasses antigen-specific Ig elution and concentration and therefore minimises antibody loss and simplifies sample preparation. Because of its high sensitivity, this technique permits clonal analysis of samples containing small quantities of Ig. We studied Ig anti-tetanus toxoid (TT) and IgG in sera from several sources. The resulting oligoclonal Ig anti-TT patterns contrasted with the polyclonal patterns of total IgG visualised after 2D-PAGE analysis of the respective serum samples.
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Département de Médecine, Centre Hospitalier Universitaire Vaudois, Lausanne, Switzerland.
Tissot
J D
JD
Schneider
P
P
Duchosal
M A
MA
D000818:Animals; D000916:Antibody Diversity; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood; D007136:Immunoglobulins / Q000032:analysis / Q000276:immunology / Q000302:isolation & purification*; D051379:Mice; D016513:Mice, SCID; D013745:Tetanus Toxoid / Q000276:immunology
D007074:Immunoglobulin G; D007136:Immunoglobulins; D013745:Tetanus Toxoid