Abstract
Rabbit exudate-derived PMN were homogenized and the cell membranes isolated on a two-phase aqueous system. Glycoproteins were extracted from cell membranes with lithium diiodosalicylate. SDS polyacrylamide gel electrophoretic analysis showed a consistent pattern of three major glycoprotein entities. Cells radioiodinated supravitally showed most of the radioactivity associated with larger glycoprotein entities whereas PMN membranes radiolabeled after isolation yielded a single major peak of radioactivity associated with a much smaller protein entity. Heterologous antisera against rabbit PMN, PMN membranes, and membrane glycoproteins were all cytotoxic for PMN in the presence of complement, and all bound to the PMN surface as demonstrated with immunocolloidal gold on electron microscopy. The data suggest that one or more glycoprotein entities are membrane-associated ectoglycoproteins which can be radiolabeled supravitally.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Goldstein, I.M., H.B. Kaplan, A. Radin, andM. Frosch. 1976. Independent effects of IgG and complement upon human polymorphonuclear leukocyte function.J. Immunol. 117:1282.
Stossel, T.P. 1975. Phagocytosis: Recognition and ingestion.In Neutrophil Physiology and Pathology. J.R. Humbert, P.A. Miescher, and E.R. Jaffé, editors. Grune and Stratton, New York. p. 93.
Scribner, D.J., andD. Fahrney. 1976. Neutrophil receptors for IgG and complement: Their roles in attachment and ingestion phases of phagocytosis.J. Immunol. 116:892.
Ehlenberger, A.G., andV. Nussenzweig. 1977. The role of membrane receptors for C3b and C3d in phagocytosis.J. Exp. Med. 145:357.
Henson, P.M. 1968. The adherence of leukocytes and platelets induced by fixed antibody and complement.Immunology 16:107.
Lay, W.H., andV. Nussenzweig. 1968. Receptors for complement on leukocytes.J. Exp. Med. 128:991.
Hawkins, D. 1977. Neutrophilic leukocytes in immunologic reactions in vitro. IV. The effect of trypsin.Inflammation 2:27.
Hawkins, D., andS. Peeters. 1971. The response of polymorphonuclear leukocytes to immune complexes in vitro.Lab. Invest. 24:483.
Marchalonis, J.J., R.E. Cone, andV. Santer. 1971. Enzymatic iodination. A probe for accessible surface proteins of normal and neoplastic lymphocytes.Biochem. J. 124:921.
Brunette, D.M., andJ.E. Till. 1971. A rapid method for the isolation of L-cell surface membranes using an aqueous two phase polymer system.J. Membr. Biol. 5:215.
McConahey, P., andF.J. Dixon. 1966. A method of trace iodination of proteins for immunologic studies.Int. Arch. Allergy 29:185.
Fishman, W.H., B. Springer, andR. Brunetti. 1948. Application of an improved glucuronidase assay method to the study of human bloodβ-glucuronidase.J. Biol. Chem. 173:449.
DePierre, J.W., andM.L. Karnovsky. 1974. Ecto-enzyme of granulocytes: 5′- nucleotidase.Science 183:1096.
Marchesi, V.T., andE.P. Andrews. 1971. Glycoproteins: Isolation from cell membranes with lithium diiodosalicylate.Science 174:1247.
Weber, K., andM. Osborne. 1969. The reliability of molecular weight determinations by dodecyl sulfate polyacrylamide gel electrophoresis.J. Biol. Chem. 244:4406.
Kapitany, E.J., andR.A. Zebrowsky. 1973. A high resolution PAS stain for polyacrylamide gel electrophoresis.Anal. Biochem. 56:361.
Hawkins, D. 1971. Biopolymer membrane: a model system for the study of the neutrophilic response to immune complexes.J. Immunol. 107:344.
Hawkins, D. 1972. Neutrophilic leukocytes in immunologic reactions: Evidence for the selective release of lysosomal constituents.J. Immunol. 108:310.
Avrameas, S., andT. Temynck. 1969. The cross-linking of proteins with glutaraldehyde and its use for the preparation of immunoabsorbents.Immunochemistry 6:53.
Hirsch, J.G., andM.E. Fedorko. 1968. Ultrastructure of human leukocytes after simultaneous fixation with glutaraldehyde and osmium tetra-oxide and “post-fixation” in uranylacetate.J. Cell Biol. 38:615.
Hawkins, D. 1974. The effect of lectins on lysosomal enzyme release from neutrophilic leukocytes.J. Immunol. 113:1864.
Horisberger, M., J. Rosset, andH. Bauer. 1975. Colloidal gold granules as markers for cell surface receptors in the scanning electron microscope.Experientia 31:1147.
Hawkins, D., andM. Sauvé. 1977. Antibody inhibition of neutrophil C3 receptor function.Clin. Res. 25:696A
Slayter, H.S., andJ.F. Codington. 1973. Size and configuration of glycoprotein fragments cleaved from tumor cells by proteolysis.J. Biol. Chem. 248:3405.
Marchesi, V.T., R.L. Jackson, J.P. Segrest, andI. Kahane. 1973. Molecular features of the major glycoprotein of the human erythrocyte membrane.Fed. Proc. 38:1833.
Nachman, R.L., A. Hubbard, andB. Ferris. 1973. Iodination of the human platelet membrane. Studies of the major surface glycoprotein.J. Biol. Chem. 248:2928.
Woodin, A.M., andA.A. Wieneke. 1966. Composition and properties of a cellmembrane fraction from the polymorphonuclear leukocyte.Biochem. J. 99:493.
Ballou, B., D.J. McKean, E.F. Freedlender, andO. Smithies. 1976. HLA membrane antigens: sequencing by intrinsic radioactivity.Proc. Natl. Acad. Sci. U.S.A. 73:4487.
Author information
Authors and Affiliations
Additional information
This study was supported by a grant from the Medical Research Council of Canada.
Rights and permissions
About this article
Cite this article
Hawkins, D., Sauvé, M. Neutrophilic leukocyte membrane proteins. Inflammation 3, 87–101 (1978). https://doi.org/10.1007/BF00917324
Issue Date:
DOI: https://doi.org/10.1007/BF00917324