Separation and Partial Purification of Plasma-Membrane Fragments from Ehrlich Ascites Carcinoma Microsomes

Kamat, V.B.; Wallach, Donald F. Hoelzl

doi:10.1126/science.148.3675.1343

Cited by 155 publications

(58 citation statements)

References 8 publications

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“…We have obtained an approximate value for the relative refractive index, m, in 0.001 M Tris-HC1 (ph 8.2) by means of Equation 8 as follows: n2 was determined as described above; nl was taken to be equal to no = 1.334, the refractive index of 0.001 i Tris-HC1, since the vesicles had been osmotically "shocked" in this medium during preparation (5,6). /3 was evaluated from the relationship (15):…”

Section: B Variation Of a With ~K Tmentioning

confidence: 99%

“…The crude PM fraction was then separated into a low density component (p = 1.025) and the purified PM component (p = 1.045) by ultracentrifugal equilibration in Ficoll gradients as described before (5,6). Prior to turbidimetric measurements it is necessary to remove the membrane components from Ficoll, since this substance has an appreciable turbidity of its own.…”

Section: Preparation Of Membrane Componentsmentioning

confidence: 99%

“…Ehrlich ascites carcinoma microsomes were separated as described previously (5,6) into a crude "plasma membrane" (PM) fraction and an "endoplasmic reticulum" (ER)~ibosome fraction by use of discontinuous Ficoll density gradients containing 0.001 M Mg 2+ and 0.001 M Tris-tICl (pH 8.6).…”

Section: Preparation Of Membrane Componentsmentioning

confidence: 99%

“…The membrane components of Ehrlich ascites carcinoma (EAC) microsomes are fluid-filled vesicles arising primarily from the disrupted endoplasmic reticulum and the fragmented plasma membrane of these cells (1)(2)(3)(4)(5). They can be fractionated by ultracentrifugal methods into four distinct membrane classes (3,5,6): namely, (1) "rough" endoplasmic reticulum; (2) vesicles--here designated ER--arising primarily from the "smooth" endoplasmic reticulum; (3) vesicles--here designated PM--originating primarily from the plasma membrane of the intact cell; and (4) particles, of unknown origin, physiochemically related to--but immunologically distinct from --PM and separated from the latter by virtue of their lower density.…”

mentioning

confidence: 99%

“…They can be fractionated by ultracentrifugal methods into four distinct membrane classes (3,5,6): namely, (1) "rough" endoplasmic reticulum; (2) vesicles--here designated ER--arising primarily from the "smooth" endoplasmic reticulum; (3) vesicles--here designated PM--originating primarily from the plasma membrane of the intact cell; and (4) particles, of unknown origin, physiochemically related to--but immunologically distinct from --PM and separated from the latter by virtue of their lower density. These particles are referred to here as the density (P) 1.025 component.…”

mentioning

confidence: 99%

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Physicochemical Differences Between Fragments of Plasma Membrane and Endoplasmic Reticulum

Wallach

Kamat

Gail

1966

The Journal of Cell Biology

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Specific turbidities, densities, and refractive indices of fragments of plasma membrane (PM) and endoplasmic reticulum (ER) from Ehrlich ascites carcinoma have been measured. A spherical shell model of specified dimensions and refractive index was established for PM fragments. The ionic composition of the dispersion medium was varied systematically. Increases in I'/2 caused increases in the turbidity of both PM and ER suspensions, the greatest effects being observed with Ca ~-and Mg 2+. In the case of PM this effect is attributable mainly to aggregation, whereas "structural" changes account for most of the turbidity increase with ER. The pH was also varied systematically to obtain pH-density and turbidity profiles and to establish the isoelectric pH of the two membrane types (PM--3.6; ER---4.35). Turbidity was maximum at "isoelectric" pH, which corresponds in each case to the region of minimum charge on the particle surfaces. Both PM and ER show large increases of density at the "isoelectric" pH, but only ER shows substantial structurally based turbidity increase under these conditions. Both PM and ER show operation of electrostatic attractions near "isoelectric" pH. PM has been shown to have ionically distinctive inner and outer surfaces while ER shows no such dissymmetry. The necessary theoretical background for interpretation of turbidity and density measurements is included, as well as a discussion of the limitations of our conclusions and the biological importance of our results.

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Section: B Variation Of a With ~K Tmentioning

confidence: 99%

Section: Preparation Of Membrane Componentsmentioning

confidence: 99%

Section: Preparation Of Membrane Componentsmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Physicochemical Differences Between Fragments of Plasma Membrane and Endoplasmic Reticulum

Wallach

Kamat

Gail

1966

The Journal of Cell Biology

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Distribution of adenosine triphosphatase activity in the developing brain of the chick embryo

Šťastný

Fröhlich

Chmelík

1969

Developmental Psychobiology

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The development and distribution of +‐K+‐stimulated and Mg2+‐dependent ATPase activity in different parts of the chick embryo brain were investigated. Maximal changes in the Na+‐K+‐stimulated ATPase activity in the hemispheres and optic lobes took place between day 15 and 17 and in the cerebellum and brain stem between day 17 and 19 of incubation. Most of this activity was localized in the membranous fraction of the chick embryo brain. An attempt was made to correlate the rapid rise in the activity of this enzyme with the development of electrical properties of the chick embryo brain.

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Helix pomatia A hemagglutinin: selectivity of binding to lymphocyte surface glycoproteins on T cells and certain B cells

et al. 1978

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Human and mouse lymphocytes were surface-labeled by lactoperoxidase-catalyzed iodination, or by galactose oxidase oxidation followed by reduction with tritiated sodium borohydride. The labeled cells were lysed with Nonidet P-40. Proteins binding to Helix pomatia A hemagglutinin (HP) were isolated by affinity chromatography on HP-Sepharose and analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis and autoradiography. A major cell surface glycoprotein (apparent mol. wt. 150 000, using reducing conditions) on human lymphocytes was responsible for almost all binding of HP. This protein was present on normal and malignant thymus-derived lymphocytes, e.g. thymocytes, blood T cells and T leukemia cell lines. It was also found on chronic lymphocytic leukemia cells, one null cell leukemia line, one unidentified leukemia line, one lymphoblastoid cell line of B origin and on one stem cell lymphoma line. In contrast, this protein was not found on various B cells at different steps of differentiation, e.g. four B lymphoma lines or one myeloma line. It was also absent from a histiocytic leukemia line. However, two of the four B lymphoma lines and the myeloma line had another HP-binding surface glycoprotein (mol. wt. 200 000) instead of the 150 000 protein. Studies of mouse lymphocytes similarly showed that thymus-derived lymphocytes (normal and malignant) but not normal adult B cells expressed a major HP-binding surface glycoprotein of apparent mol. wt. 130 000 (reducing conditions).

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Separation and Partial Purification of Plasma-Membrane Fragments from Ehrlich Ascites Carcinoma Microsomes

Cited by 155 publications

References 8 publications

Physicochemical Differences Between Fragments of Plasma Membrane and Endoplasmic Reticulum

Physicochemical Differences Between Fragments of Plasma Membrane and Endoplasmic Reticulum

Distribution of adenosine triphosphatase activity in the developing brain of the chick embryo

Helix pomatia A hemagglutinin: selectivity of binding to lymphocyte surface glycoproteins on T cells and certain B cells

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