Abstract
1. A method was developed which allows the rapid preparation of pure plasma membranes in high yield from cultured fibroblasts. 2. Cells are lysed in hypo-osmotic borate/EDTA and, after differential centrifugation, the membranes collected by centrifugation on a sucrose barrier. 3. Electron microscopy of the isolated material shows large membrane vesicles essentially free from contaminating organelles. 4. There is no detectable activity of the endoplasmic-reticulum enzyme marker, NADH2--lipoamide oxidoreductase (EC 1.6.4.3), and that of succinate dehydrogenase (EC 1.3.99.1), a marker for mitochondria, is substantially decreased. Chemical compositions are in good agreement with previous observations. 5. This study confirms the usefulness of applied isotopic markers for isolating plasma membranes.
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- BLIX G., LINDBERG E., ODIN L., WERNER I. Studies on sialic acids. Acta Soc Med Ups. 1956 Jun 30;61(1-2):1–25. [PubMed] [Google Scholar]
- COOK G. M., HEARD D. H., SEAMAN G. V. A sialomucopeptide liberated by trypsin from the human erythrocyte. Nature. 1960 Dec 17;188:1011–1012. doi: 10.1038/1881011a0. [DOI] [PubMed] [Google Scholar]
- Chang K. J., Bennett V., Cuatrecasas P. Membrane receptors as general markers for plasma membrane isolation procedures. The use of 125-I-labeled wheat germ agglutinin, insulin, and cholera toxin. J Biol Chem. 1975 Jan 25;250(2):488–500. [PubMed] [Google Scholar]
- Critchley D. R., Wyke J. A., Hynes R. O. Cell surface and metabolic labelling of the proteins of normal and transformed chicken cells. Biochim Biophys Acta. 1976 Jun 17;436(2):335–352. doi: 10.1016/0005-2736(76)90198-x. [DOI] [PubMed] [Google Scholar]
- Fairbanks G., Steck T. L., Wallach D. F. Electrophoretic analysis of the major polypeptides of the human erythrocyte membrane. Biochemistry. 1971 Jun 22;10(13):2606–2617. doi: 10.1021/bi00789a030. [DOI] [PubMed] [Google Scholar]
- Ferber E., Resch K., Wallach D. F., Imm W. Isolation and characterization of lymphocyte plasma membranes. Biochim Biophys Acta. 1972 May 9;266(2):494–504. doi: 10.1016/0005-2736(72)90105-8. [DOI] [PubMed] [Google Scholar]
- Gahmberg C. G., Simons K. Isolation of plasma membrane fragments from BHK21 cells. Acta Pathol Microbiol Scand B Microbiol Immunol. 1970;78(2):176–182. doi: 10.1111/j.1699-0463.1970.tb04284.x. [DOI] [PubMed] [Google Scholar]
- Juliano R. L., Gagalang E. The isolation and characterization of the plasma membrane of cultured chinese hamster ovary cells. Can J Biochem. 1975 Aug;53(8):914–920. doi: 10.1139/o75-126. [DOI] [PubMed] [Google Scholar]
- 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]
- LANGLEY O. K., AMBROSE E. J. ISOLATION OF A MUCOPEPTIDE FROM THE SURFACE OF EHRLICH ASCITES TUMOUR CELLS. Nature. 1964 Oct 3;204:53–54. doi: 10.1038/204053a0. [DOI] [PubMed] [Google Scholar]
- 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]
- Liao T. H., Gallop P. M., Blumenfeld O. O. Modification of sialyl residues of sialoglycoprotein(s) of the human erythrocyte surface. J Biol Chem. 1973 Dec 10;248(23):8247–8253. [PubMed] [Google Scholar]
- Lloyd C. W., Cook G. M. On the mechanism of the increased aggregation by neuraminidase of 16C malignant rat dermal fibroblasts in vitro. J Cell Sci. 1974 Aug;15(3):575–590. doi: 10.1242/jcs.15.3.575. [DOI] [PubMed] [Google Scholar]
- McCollester D. L. Isolation of meth A cell surface membranes possessing tumor-specific transplantation antigen activity. Cancer Res. 1970 Dec;30(12):2832–2837. [PubMed] [Google Scholar]
- Novogrodsky A., Katchalski E. Membrane site modified on induction of the transformation of lymphocytes by periodate. Proc Natl Acad Sci U S A. 1972 Nov;69(11):3207–3210. doi: 10.1073/pnas.69.11.3207. [DOI] [PMC free article] [PubMed] [Google Scholar]
- PORTEOUS J. W., CLARK B. THE ISOLATION AND CHARACTERIZATION OF SUBCELLULAR COMPONENTS OF THE EPITHELIAL CELLS OF RABBIT SMALL INTESTINE. Biochem J. 1965 Jul;96:159–171. doi: 10.1042/bj0960159. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Perdue J. F., Sneider J. The isolation and characterization of the plasma membrane from chick embryo fibroblasts. Biochim Biophys Acta. 1970;196(2):125–140. doi: 10.1016/0005-2736(70)90001-5. [DOI] [PubMed] [Google Scholar]
- Persijn J. P., van der Slik W., Kramer K., de Ruijter C. A. A new method for the determination of serum nucleotidase. Z Klin Chem Klin Biochem. 1968 Sep;6(5):441–446. doi: 10.1515/cclm.1968.6.5.441. [DOI] [PubMed] [Google Scholar]
- Renkonen O., Gahmberg C. G., Simons K., Käriäinen L. The lipids of the plasma membranes and endoplasmic reticulum from cultured baby hamster kidney cells (BHK21). Biochim Biophys Acta. 1972 Jan 17;255(1):66–78. doi: 10.1016/0005-2736(72)90008-9. [DOI] [PubMed] [Google Scholar]
- Rittenhouse-Simmons S., Deykin D. Isolation of membranes from normal and thrombin-treated gel-filtered platelets using a lectin marker. Biochim Biophys Acta. 1976 Apr 5;426(4):688–696. doi: 10.1016/0005-2736(76)90133-4. [DOI] [PubMed] [Google Scholar]
- Rouser G., Siakotos A. N., Fleischer S. Quantitative analysis of phospholipids by thin-layer chromatography and phosphorus analysis of spots. Lipids. 1966 Jan;1(1):85–86. doi: 10.1007/BF02668129. [DOI] [PubMed] [Google Scholar]
- Spurr A. R. A low-viscosity epoxy resin embedding medium for electron microscopy. J Ultrastruct Res. 1969 Jan;26(1):31–43. doi: 10.1016/s0022-5320(69)90033-1. [DOI] [PubMed] [Google Scholar]
- Thomas D. B., Winzler R. J. Structural studies on human erythrocyte glycoproteins. Alkali-labile oligosaccharides. J Biol Chem. 1969 Nov 10;244(21):5943–5946. [PubMed] [Google Scholar]
- Van Lenten L., Ashwell G. Studies on the chemical and enzymatic modification of glycoproteins. A general method for the tritiation of sialic acid-containing glycoproteins. J Biol Chem. 1971 Mar 25;246(6):1889–1894. [PubMed] [Google Scholar]
- Warley A., Cook G. M. The isolation and characterization of plasma membranes from normal and leukaemic cells of mice. Biochim Biophys Acta. 1973 Sep 27;323(1):55–68. doi: 10.1016/0005-2736(73)90431-8. [DOI] [PubMed] [Google Scholar]
- Winzler R. J., Harris E. D., Pekas D. J., Johnson C. A., Weber P. Studies on glycopeptides released by trypsin from intact human erythrocytes. Biochemistry. 1967 Jul;6(7):2195–2202. doi: 10.1021/bi00859a042. [DOI] [PubMed] [Google Scholar]
- ZLATKIS A., ZAK B., BOYLE A. J. A new method for the direct determination of serum cholesterol. J Lab Clin Med. 1953 Mar;41(3):486–492. [PubMed] [Google Scholar]

