The Isolation of the Surface Membranes of Animal Cells: A Survey

Warren, Leonard; Glick, Mary Catherine

doi:10.1007/978-1-4684-3327-2_4

Cited by 18 publications

(5 citation statements)

References 91 publications

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“…Better yields and membranes of comparable or higher purity (~ 14-fold) were obtained by simplifying the original Warren procedure (FMA II). FMA membranes prepared by either procedure displayed fibrillar material and many ribosome-like particles adsorbed to the cytoplasmic face, as reported by others using different tissue culture cells (28,63). These particles did not appear to be attached to the plasma membrane in the intact cell (20) and exposure of the isolated membranes to a high salt concentration (0.5 M KC1) released most of them (1).…”

Section: Membrane-enriched Preparationssupporting

confidence: 57%

“…Methods published by Warren and colleagues (63,64) employing Zn § fluorescein mercuric acetate (FMA), and Tris-HC1 at pH 7.4 and 8.0 as membrane-"stabilizing" agents were followed and found to be time consuming and quite variable in yield and purification, as assessed by TCA-insolubte ~2~I per milligram protein.…”

Section: Isolation Of L Cell Plasma Membranementioning

confidence: 99%

“…The L cell divides rapidly so large numbers can be easily obtained (108 in 500 ml), and it will phagocytose polystyrene latex particles, thus internalizing a large proportion of its plasma membrane in a short time (17). In addition, there are published reports on the isolation of L-cell plasma membrane (63,64).…”

mentioning

confidence: 99%

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Externally disposed plasma membrane proteins. I. Enzymatic iodination of mouse L cells.

Hubbard

Cohn

1975

The Journal of Cell Biology

346

162

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The enzymatic iodination technique has been utilized in a study of the externally disposed membrane proteins of the mouse L cell. lodination of cells in suspension results in lactoperoxidase-specific iodide incorporation with no loss of cell viability under the conditions employed, less than 3% lipid labeling, and more than 90% of the labeled species identifiable as monoiodotyrosine. 90% of the incorporated label is localized to the cell surface by electron microscope autoradiography, with 5-10% in the centrosphere region and postulated to represent pinocytic vesicles. Sodium dodecylsulfate-polyacrylamide gels of solubilized L-cell proteins reveals five to six labeled peaks ranging from 50,000 to 200,000 daltons. Increased resolution by use of gradient slab gels reveals 15-20 radioactive bands. Over 60% of the label resides in approximately nine polypeptides of 80,000 to 150,000 daltons. Various controls indicate that the labeling pattern reflects endogenous membrane proteins, not serum components. The incorporated 125I, cholesterol, and one plasma membrane enzyme marker, alkaline phosphodiesterase I, are purified in parallel when plasma membranes are isolated from intact, iodinated L cells. The labeled components present in a plasma membrane-rich fraction from iodinated cells are identical to those of the total cell, with a 10-to 20-fold enrichment in specific activity of each radioactive peak in the membrane.

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Section: Membrane-enriched Preparationssupporting

confidence: 57%

Section: Isolation Of L Cell Plasma Membranementioning

confidence: 99%

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Externally disposed plasma membrane proteins. I. Enzymatic iodination of mouse L cells.

Hubbard

Cohn

1975

The Journal of Cell Biology

346

162

View full text Add to dashboard Cite

show abstract

“…The procedure was monitored in the phase contrast microscope, and after gentle homogenization in a Dounce with a loose-fitting pestle the membrane can be observed coming off the cell. Sucrose gradients were used to separate the membranes from cell particulates, nuclei, and remaining whole cells as described (Warren and Glick, 1969). The surface membranes prepared by this method were identifiable morphologically 30% sucrose and visualized in a Zeiss phase contrast microscope.…”

Section: Preparation Of Surface Membranesmentioning

confidence: 99%

Turnover of the cystic fibrosis transmembrane conductance regulator (CFTR): Slow degradation of wild-type and ΔF508 CFTR in surface membrane preparations of immortalized airway epithelial cells

Wei

Eisman

et al. 1996

J. Cell. Physiol.

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The protein product of the cystic fibrosis (CF) gene, termed the cystic fibrosis transmembrane conductance regulator (CFTR), is known to function as an apical chloride channel at the surface of airway epithelial cells. It has been proposed that CFTR has additional intracellular functions and that there is altered processing of mutant forms. In examining these functions we found a stable form of CFTR with slow turnover in surface membrane preparations from CF and non-CF immortalized airway epithelial cell lines. The methods used to study the turnover of CFTR were pulse/chase experiments utilizing saturation labeling of [35S] Met with chase periods of 5-24 h in the presence of 8 mM Met and cell fractionation techniques. Preparations of morphologically identifiable surface membranes were compared to total cell membrane preparations containing intracellular membranes. Surface membrane CFTR had lower turnover defined by pulse/chase ratios than that of the total cell membrane preparations. Moreover, mutant CFTR was stable in the surface membrane fraction with little degradation even after a 24 h chase, whereas wild-type CFTR had a higher pulse/chase ratio at 24 h. In the presence of 50 microM castanospermine, which is an inhibitor of processing alpha-glucosidases, a more rapid turnover of mutant CFTR was found in the total cell membrane preparation, whereas wild-type CFTR had a lower response. The results are compatible with a pool of CFTR in or near the surface membranes which has an altered turnover in CF and a glycosylation-dependent alteration in the processing of mutant CFTR.

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“…Πρόσθετα τά μικρού μήκους αλυσίδας FA έχουν μικρότερο εμβαδόν καί ευκολία γιά περιστροφική κίνηση. 13 Στίς επιφανειακές 1 δμως μεμβράνες πού περιέχουν μιά ποικιλία λιπιδικών τάξεων καί σέ κά θε τάξη αντιστοιχεί ένα μεγάλο καί συναρπαστικό φάσμα FA,χρειάστηκε νά γίνουν μερικές χρή σιμες γενικεύσεις. "Ετσι γιά κάθε τάξη μεμβρανικοΰ λιπιδίου καί γιά κάθε ϋερμοκρασία ή μεμβρανική ρευστότητα είναι μεγαλύτερη γιά μικρό μήκος ύδρογονανθρακικής αλυσίδας, καί γιά περισσότερο ακόρεστα άκυλο-παράγωγα.…”

unclassified

Crithidia Fasciculata: Ενα Συστημα Για Την Μελετη Της Δυναμικης Συμπεριφορας Των Επιφανειακων Μεμβρανων

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The Isolation of the Surface Membranes of Animal Cells: A Survey

Cited by 18 publications

References 91 publications

Externally disposed plasma membrane proteins. I. Enzymatic iodination of mouse L cells.

Externally disposed plasma membrane proteins. I. Enzymatic iodination of mouse L cells.

Turnover of the cystic fibrosis transmembrane conductance regulator (CFTR): Slow degradation of wild-type and ΔF508 CFTR in surface membrane preparations of immortalized airway epithelial cells

Crithidia Fasciculata: Ενα Συστημα Για Την Μελετη Της Δυναμικης Συμπεριφορας Των Επιφανειακων Μεμβρανων

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