Microisolation of DABITC-derivatized protein by gel electrophoresis: application to the purification of antibody heavy and light chains.
Proteins and polypeptides are derivatized with dimethylaminoazobenzene isothiocyanate (DABITC) before their separation on sodium dodecyl sulfate-polyacrylamide gels. DABITC-derivatized proteins are detected visually in the picomole range without further staining and destaining procedures. The recovered colored protein can also be used for direct sequence determinations. This method was applied to purify heavy and light chains of murine hybridoma-derived antibodies. Starting with ammonium sulfate-precipitated protein mixture, this method allows, in a one-step procedure, the isolation of pure light and heavy chains suitable for automatic sequencing.
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D006825:Hybridomas / Q000276:immunology; D007143:Immunoglobulin Heavy Chains / Q000302:isolation & purification*; D007147:Immunoglobulin Light Chains / Q000302:isolation & purification*; D007202:Indicators and Reagents; D017879:Isothiocyanates; D051379:Mice; D013861:Thiocyanates; D004124:p-Dimethylaminoazobenzene / Q000031:analogs & derivatives
D000906:Antibodies; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007202:Indicators and Reagents; D017879:Isothiocyanates; D013861:Thiocyanates; C012926:4-(N,N-dimethylaminoazobenzene)-4'-isothiocyanate; D004124:p-Dimethylaminoazobenzene