Separation of antibody-antigen complexes by capillary zone electrophoresis, isoelectric focusing and high-performance size-exclusion chromatography.
The separation of antibody-antigen complexes by free-solution capillary zone electrophoresis (CZE) has been demonstrated. The antigen, a monoclonal antibody specific for the antigen, and the complex were well resolved. The entire separation was achieved in less than 10 min using on-column UV detection. The pI values for the three species were estimated separately by isoelectric focusing (IEF) experiments on polyacrylamide gels. Reasonably good agreement was found between the relative migration times measured by CZE and the pI values. Both IEF and high-performance size-exclusion chromatography of the antibody-antigen mixtures confirmed the formation of the complexes observed by CZE. This study demonstrates the utility of CZE as a new and complementary technique for the characterization of antibody-antigen complexes.
Nielsen
R G
RG
Lilly Research Laboratories, Eli Lilly & Company, Indianapolis, IN 46285.
Rickard
E C
EC
Santa
P F
PF
Sharknas
D A
DA
Sittampalam
G S
GS
D000911:Antibodies, Monoclonal / Q000276:immunology / Q000302:isolation & purification; D000936:Antigen-Antibody Complex / Q000302:isolation & purification*; D000941:Antigens / Q000276:immunology / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid; D004586:Electrophoresis / Q000379:methods*; D013006:Growth Hormone / Q000276:immunology; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology / Q000302:isolation & purification; D007525:Isoelectric Focusing; D011994:Recombinant Proteins / Q000276:immunology
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D000941:Antigens; D007074:Immunoglobulin G; D011994:Recombinant Proteins; D013006:Growth Hormone