Skip to main content
NCBI home page
Biochemical Journal logoLink to Biochemical Journal
. 1970 Nov;120(1):133–143. doi: 10.1042/bj1200133

Preparation and characterization of the plasma membrane of pig lymphocytes

D Allan 1, M J Crumpton 1
PMCID: PMC1179577  PMID: 4321928

Abstract

Lymphocyte plasma membrane was isolated from minced pig mesenteric lymph node by differential centrifugation and by centrifuging through a sucrose density gradient. The yield of membrane was approx. 0.1% (dry wt. relative to wet wt. of lymph node). The purified material had a sucrose density of 1.14g/cm3 and consisted mainly of smooth vesicles. The membrane fraction contained, apart from protein and lipid, 59μg of carbohydrate, 11μg of sialic acid and 28μg of RNA/mg of protein; no DNA was detected. The cholesterol/phospholipid molar ratio was 1.01. Specific activities (μmol of product/h per mg of protein) of 5′-nucleotidase, succinate dehydrogenase, acid phosphatase and glucose 6-phosphatase were 10.1, 0, 0.51 and 0.30 respectively. The membrane vesicles were aggregated by an antiserum against pig lymphocytes and adsorbed the agglutinins to whole lymphocytes present in the antiserum; the membrane fraction was 28 times as effective as whole cells (on a dry wt. basis) in removing the lympho-agglutinins. Antisera against the membrane fraction agglutinated whole lymphocytes. It is concluded that the preparation represents the plasma membrane of small lymphocytes. The plasma membrane of pig thymocytes was isolated by using the same procedure. Its properties were similar to those of the lymphocyte plasma membrane.

Full text

PDF
133

Images in this article

140-2PLATE 2
on p.140-2
140-3PLATE 3
on p.140-3
140-4PLATE 4
on p.140-4
140-1PLATE 1
on p.140-1

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. BENSON J. V., Jr, PATTERSON J. A. ACCELERATED AUTOMATIC CHROMATOGRAPHIC ANALYSIS OF AMINO ACIDS ON A SPHERICAL RESIN. Anal Chem. 1965 Aug;37:1108–1110. doi: 10.1021/ac60228a008. [DOI] [PubMed] [Google Scholar]
  2. Bakerman S., Wasemiller G. Studies on structural units of human erythrocyte membrane. I. Separation, isolation, and partial characterization. Biochemistry. 1967 Apr;6(4):1100–1113. doi: 10.1021/bi00856a020. [DOI] [PubMed] [Google Scholar]
  3. Boone C. W., Ford L. E., Bond H. E., Stuart D. C., Lorenz D. Isolation of plasma membrane fragments from HeLa cells. J Cell Biol. 1969 May;41(2):378–392. doi: 10.1083/jcb.41.2.378. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bosmann H. B., Hagopian A., Eylar E. H. Cellular membranes: the isolation and characterization of the plasma and smooth membranes of HeLa cells. Arch Biochem Biophys. 1968 Oct;128(1):51–69. doi: 10.1016/0003-9861(68)90008-8. [DOI] [PubMed] [Google Scholar]
  5. Bowers W. E., de Duve C. Lysosomes in lymphoid tissue. III. Influence of various treatments of the animals on the distribution of acid hydrolases. J Cell Biol. 1967 Feb;32(2):349–364. doi: 10.1083/jcb.32.2.349. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Brittinger G., Hirschhorn R., Douglas S. D., Weissmann G. Studies on lysosomes. XI. Characterization of a hydrolase-rich fraction from human lymphocytes. J Cell Biol. 1968 May;37(2):394–411. doi: 10.1083/jcb.37.2.394. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. CRUMPTON M. J., WILKINSON J. M. AMINO ACID COMPOSITIONS OF HUMAN AND RABBIT GAMMA-GLOBULINS AND OF THE FRAGMENTS PRODUCED BY REDUCTION. Biochem J. 1963 Aug;88:228–234. doi: 10.1042/bj0880228. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Coleman R., Finean J. B. Structural and biochemical features of isolated cell surface membranes. Protoplasma. 1967;63(1):172–173. [PubMed] [Google Scholar]
  9. DODGE J. T., MITCHELL C., HANAHAN D. J. The preparation and chemical characteristics of hemoglobin-free ghosts of human erythrocytes. Arch Biochem Biophys. 1963 Jan;100:119–130. doi: 10.1016/0003-9861(63)90042-0. [DOI] [PubMed] [Google Scholar]
  10. Davidova S. Y., Shapot V. S. Liporibonucleoprotein complex as an integral part of animal cell plasma membranes. FEBS Lett. 1970 Feb 25;6(4):349–351. doi: 10.1016/0014-5793(70)80096-5. [DOI] [PubMed] [Google Scholar]
  11. Dumonde D. C., Wolstencroft R. A., Panayi G. S., Matthew M., Morley J., Howson W. T. "Lymphokines": non-antibody mediators of cellular immunity generated by lymphocyte activation. Nature. 1969 Oct 4;224(5214):38–42. doi: 10.1038/224038a0. [DOI] [PubMed] [Google Scholar]
  12. EARL D. C., KORNER A. THE ISOLATION AND PROPERTIES OF CARDIAC RIBOSOMES AND POLYSOMES. Biochem J. 1965 Mar;94:721–734. doi: 10.1042/bj0940721. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Evans W. H. Fractionation of liver plasma membranes prepared by zonal centrifugation. Biochem J. 1970 Mar;116(5):833–842. doi: 10.1042/bj1160833. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Fitzpatrick D. F., Davenport G. R., Forte L., Landon E. J. Characterization of plasma membrane proteins in mammalian kidney. I. Preparation of a membrane fraction and separation of the protein. J Biol Chem. 1969 Jul 10;244(13):3561–3569. [PubMed] [Google Scholar]
  15. GIANETTO R., DE DUVE C. Tissue fractionation studies. 4. Comparative study of the binding of acid phosphatase, beta-glucuronidase and cathepsin by rat-liver particles. Biochem J. 1955 Mar;59(3):433–438. doi: 10.1042/bj0590433. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Glick M. C., Warren L. Membranes of animal cells. 3. Amino acid incorporation by isolated surface membranes. Proc Natl Acad Sci U S A. 1969 Jun;63(2):563–570. doi: 10.1073/pnas.63.2.563. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hirsch J. G., Fedorko M. E. Ultrastructure of human leukocytes after simultaneous fixation with glutaraldehyde and osmium tetroxide and "postfixation" in uranyl acetate. J Cell Biol. 1968 Sep;38(3):615–627. doi: 10.1083/jcb.38.3.615. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. KLENK E., UHLENBRUCK G. Uber ein neuraminsäurehaltiges Mucoproteid aus Rindererythrocytenstroma. Hoppe Seylers Z Physiol Chem. 1958;311(4-6):227–233. [PubMed] [Google Scholar]
  19. Kahan B. D., Reisfeld R. A. Transplantaton antigens. Science. 1969 May 2;164(3879):514–521. doi: 10.1126/science.164.3879.514. [DOI] [PubMed] [Google Scholar]
  20. Kopaczyk K., Perdue J., Green D. E. The relation of structural and catalytic protein in the mitochondrial electron transfer chain. Arch Biochem Biophys. 1966 Jul;115(1):215–225. doi: 10.1016/s0003-9861(66)81060-3. [DOI] [PubMed] [Google Scholar]
  21. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  22. Michell R. H., Hawthorne J. N. The site of diphosphoinositide synthesis in rat liver. Biochem Biophys Res Commun. 1965 Nov 22;21(4):333–338. doi: 10.1016/0006-291x(65)90198-1. [DOI] [PubMed] [Google Scholar]
  23. Raff M. C., Sternberg M., Taylor R. B. Immunoglobulin determinants on the surface of mouse lymphoid cells. Nature. 1970 Feb 7;225(5232):553–554. doi: 10.1038/225553a0. [DOI] [PubMed] [Google Scholar]
  24. SWANSON M. A. Phosphatases of liver. I. Glucose-6-phosphatase. J Biol Chem. 1950 Jun;184(2):647–659. [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES