title: Purification and immunochemical characterization of a natural human polyreactive monoclonal IgM antibody.
abstract: In vitro and in vivo experiments to explain the function of natural polyreactive antibodies, usually of the IgM isotype, require large amounts of purified antibodies. We have developed a two-step purification procedure using a human natural polyreactive monoclonal IgM antibody (CB03). This combines hydrophobic interaction chromatography on phenyl-Superose and gel filtration over Superose 12 and readily permits scaling-up to isolate mg to g amounts of antibody. Retention of the CB03 antibody during gel filtration by precipitation and interaction with the gel matrix was overcome by the addition of 10 mM 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonate. The yield of purified antibody was 34% and Fab fragments were obtained from the purified CB03 antibody by hot tryptic digestion (yield, 68% of theoretical amount). In an enzyme-linked immunosorbent assay, Fab and complete antibody had similar reaction patterns with different antigens.
authors.affiliation: Department of Medical Immunology, Medical School (Charité), Humboldt University of Berlin, Germany.
authors.forename: D
authors.initials: D
authors.lastname: Roggenbuck
authors.forename: U
authors.initials: U
authors.lastname: Marx
authors.forename: S T
authors.initials: ST
authors.lastname: Kiessig
authors.forename: G
authors.initials: G
authors.lastname: Schoenherr
authors.forename: S
authors.initials: S
authors.lastname: Jahn
authors.forename: T
authors.initials: T
authors.lastname: Porstmann
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000302:isolation & purification; D000918:Antibody Specificity; D002845:Chromatography; D006801:Humans; D006825:Hybridomas; D007140:Immunoglobulin Fab Fragments / Q000276:immunology / Q000302:isolation & purification; D007075:Immunoglobulin M / Q000276:immunology* / Q000302:isolation & purification; D008214:Lymphocytes / Q000276:immunology; D013154:Spleen / Q000166:cytology; D013921:Thrombocytopenia / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D007075:Immunoglobulin M