Effects of chemical modification of antibodies on their clearance from the circulation. Addition of simple aliphatic compounds by reductive alkylation and carbodiimide-promoted amide formation.
Anti-hapten antibodies from the ascitic fluid of inbred mice were purified by immunoadsorption and characterized immunochemically for in vivo studies of their plasma clearance rates and organ distributions after chemical modification. Following sodium borohydride-promoted reductive methylation and carbodiimide-promoted amide linkage of glycine and several other simple aliphatic compounds, the antibody populations were recharacterized, radiolabeled, and introduced intravenously into syngeneic animals. Using double radioiodine labels, it was possible to show that stoichiometric combinations of additive and chemical reactant did not alter antibody survival time in the circulation or antigen-binding capabilities. However, modifications involving excess reagent (carbodiimide or sodium borohydride) resulted in significant decreases in both circulatory longevity and ligand binding capacities. Excess carbodiimide treatments resulted in immunoglobulin cross-linkage which could be detected by molecular sieve chromatography. Increased kidney localization found with carbodiimide-treated antibodies was apparently due to trapping of the cross-linked aggregates. Elevated clearance of highly methylated antibodies could not be attributed to a single organ. However, sodium borohydride-catalyzed reductive methylation resulted in antibody populations which were localized primarily in the liver and spleen. Results are evaluated in terms of the concept, developed in this paper, of essential groups for the circulatory longevity of glycoproteins.
D000478:Alkylation; D000577:Amides; D000818:Animals; D000937:Antigen-Antibody Reactions; D001894:Borohydrides; D002234:Carbodiimides; D005998:Glycine / Q000378:metabolism; D007074:Immunoglobulin G / Q000378:metabolism; D007668:Kidney / Q000378:metabolism; D007700:Kinetics; D010084:Oxidation-Reduction; D011485:Protein Binding; D011817:Rabbits / Q000276:immunology; D013552:Swine / Q000276:immunology
D000577:Amides; D001894:Borohydrides; D002234:Carbodiimides; D007074:Immunoglobulin G; D005998:Glycine