Human monoclonal antibody recognizing liver-type aldolase B.
A human hybridoma clone (4E3) has been established by fusing lymphocytes from a lymph node taken from a breast cancer patient and human lymphoblastoid cells, LICR-LON-HMy2, by the poly(ethylene glycol) method. 4E3 has been stabilized and continued to secrete IgMk antibody into culture medium (greater than 10 micrograms/ml) for over 1 year. The following characteristics of the antigen strongly suggested that 4E3 recognizes liver-type aldolase B (EC 4.1.2.13): the Mr of the native molecule is 160,000 and that of the subunit is 40,000, and thus it has a tetrameric structure of identical subunits; the antigen is abundant in the liver and kidney of human, mouse and rabbit, and is localized by immunohistochemical methods in the cytoplasm of hepatocytes and in the proximal tubules of the kidney; the antigen is precipitable by 50-80% saturation with (NH4)2SO4; the antigen shows charge-dependent heterogeneity on DEAE-cellulose chromatography. To confirm this notion, aldolase B was purified to homogeneity from the liver of human, mouse and rabbit by phosphocellulose chromatography. During the chromatographic purification, the antigen activity as assayed by enzyme-linked immunosorbent assay (e.l.i.s.a.) was superimposed on the enzymic activity of aldolase. Furthermore, monoclonal antibody 4E3 strongly reacted with purified aldolase B in SDS/polyacrylamide-gel electrophoresis followed by Western blotting and also in e.l.i.s.a. using microplates coated with purified enzyme. The reaction between aldolase B and 4E3 activated the human complement system as assessed by the attachment of C3 to the immune complex of aldolase B and 4E3.
D000818:Animals; D000911:Antibodies, Monoclonal; D000941:Antigens / Q000302:isolation & purification; D002460:Cell Line; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005634:Fructose-Bisphosphate Aldolase / Q000276:immunology* / Q000302:isolation & purification; D006801:Humans; D007122:Immunoelectrophoresis; D007527:Isoenzymes / Q000276:immunology* / Q000302:isolation & purification; D007668:Kidney / Q000201:enzymology; D008099:Liver / Q000201:enzymology; D051379:Mice; D013347:Subcellular Fractions / Q000201:enzymology
D000911:Antibodies, Monoclonal; D000941:Antigens; D007527:Isoenzymes; D005634:Fructose-Bisphosphate Aldolase