High-performance sodium dodecyl sulfate-capillary gel electrophoresis of antibodies and antibody fragments.
High-performance sodium dodecyl sulfate-capillary gel electrophoresis (SDS-CGE) has been used to separate antibodies and their fragments according to size. Under non-reduced conditions, murine monoclonal antibodies generally show a predominant peak with five to six apparent fragment peaks. The magnitude of the apparent fragmentation is temperature-dependent and is more pronounced with rabbit, sheep and bovine immunoglobulin G's than murine monoclonal antibodies. In addition to temperature, pH and buffer also affect the fragmentation. Without heat treatment during the preparation of the SDS-antibody complexes, the observed fragments become nearly absent; however, some murine monoclonal antibodies exhibit several peaks that group near the expected migration time of an immunoglobulin G, presumably due to their anomalous interaction with sodium dodecyl sulfate. The method can also be used to monitor the progress of peptic digestions to generate murine F(ab')(2) antibody, to perform "gel-shift" assays, and to separate mouse ascitic fluid. This high-performance electrophoretic technique is suitable for quality control as well as the characterization of the antibodies under experimental conditions.
Lee
H G
HG
Product Development, Diagnostics Division, Bayer, East Walpole, MA 02032, USA. huey.lee.b@bayer.com
D000818:Animals; D000911:Antibodies, Monoclonal / Q000302:isolation & purification*; D002417:Cattle; D019075:Electrophoresis, Capillary / Q000379:methods*; D007140:Immunoglobulin Fab Fragments / Q000302:isolation & purification; D007128:Immunoglobulin Fragments / Q000302:isolation & purification*; D051379:Mice; D011817:Rabbits; D012756:Sheep; D012967:Sodium Dodecyl Sulfate; D013552:Swine
D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D007128:Immunoglobulin Fragments; D012967:Sodium Dodecyl Sulfate