Solid phase synthesis of bifunctional antibodies.
Bifunctional antibodies were prepared using the principle of solid-phase synthesis. The two Fab' fragments were chemically linked together via a bismaleimide crosslinking reagent. The F(ab')2 fragments from intact IgG were prepared using an immobilized pepsin column. Goat, mouse and human antibodies were digested completely within 4 h. The F(ab')2 fragments thus produced did not contain any IgG impurities. The Fab' fragments were produced by reducing the inter-heavy chain disulfide bonds using 2-mercaptoethylamine. The use of the solid-phase reactor in the preparation of the bifunctional antibodies eliminated many of the time-consuming separation steps between the fragmentation and conjugation steps. This procedure facilitates the automation of the bifunctional antibody preparation and the rapid optimization of reaction conditions.
DeSilva
B S
BS
Department of Chemistry, University of Kansas, Lawrence 66045, USA.
Wilson
G S
GS
D000818:Animals; D018033:Antibodies, Bispecific / Q000096:biosynthesis* / Q000737:chemistry; D002850:Chromatography, Gel; D003432:Cross-Linking Reagents; D004800:Enzymes, Immobilized; D006041:Goats; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis / Q000737:chemistry; D007074:Immunoglobulin G / Q000096:biosynthesis* / Q000737:chemistry; D008624:Mercaptoethylamines; D051379:Mice; D010084:Oxidation-Reduction; D010434:Pepsin A
D018033:Antibodies, Bispecific; D003432:Cross-Linking Reagents; D004800:Enzymes, Immobilized; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D008624:Mercaptoethylamines; D010434:Pepsin A