Microtubules in mouse embryo fibro blasts extracted with Triton X-100

Bershadsky, Alexander D.; Gelfand, Vladimir I.; Svitkina, Tatyana; Tint, I.S.

doi:10.1016/0309-1651(78)90093-0

Cited by 61 publications

(22 citation statements)

References 12 publications

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“…This method preserves microtubule and microfilament networks (Bershadsky et al, 1978). As observed by scanning electron microscopy, the cytoskeleton was well-developed in control ( Fig.…”

Section: Analysis Of the Cytoskeletal Network In Ifn-treated Y-1 Cellmentioning

confidence: 78%

“…(105) were grown for 24 h on glass coverslips in culture dishes. After 24 h treatment with either IFN, cholera toxin or serum-free medium, the cells were washed and extracted for 5 min at 37 °C with 1% Triton X-100 (Bershadsky et al, 1978). Glutaraldehyde-fixed (1%) cells were dehydrated in graded ethanol series and then acetone, dried using a CO2 critical point method (Balzer CPD 010) and coated with 10 nm-thick Au-Pd by sputtering (Balzer SCD 020).…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Factors Involved in Interferon-induced or Cholera Toxin-induced Steroidogenesis in Y-1 Mouse Adrenal Tumour Cells

Matsuguchi¹,

Moreau²,

Rousset³

et al. 1985

Journal of General Virology

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SUMMARYIn addition to its antiviral effect, interferon, at high concentrations, stimulates steroidogenesis and provokes cell rounding in Y-1 mouse adrenal tumour cells. This stimulation was inhibited by cytochalasin B and colchicine. In contrast, dibutyryl cAMP and cholera toxin, also able to induce steroid production and cell rounding, increased steroid production even in the presence of these cytoskeleton-disrupting agents. The initial trigger for interferon or cholera toxin thus probably involves a distinct receptor organization. However, since both inducers increased cAMP synthesis in this differentiated cell line, the further metabolic steps of ketosteroid production could be the same.

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“…This method preserves microtubule and microfilament networks (Bershadsky et al, 1978). As observed by scanning electron microscopy, the cytoskeleton was well-developed in control ( Fig.…”

Section: Analysis Of the Cytoskeletal Network In Ifn-treated Y-1 Cellmentioning

confidence: 78%

Section: Methodsmentioning

confidence: 99%

Factors Involved in Interferon-induced or Cholera Toxin-induced Steroidogenesis in Y-1 Mouse Adrenal Tumour Cells

Matsuguchi¹,

Moreau²,

Rousset³

et al. 1985

Journal of General Virology

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“…From the electron microscopic observations of epoxyembedded sections, the filaments resistant to the extraction by 1% Triton X-100 were thought to represent MFs, MTs and IFs, and those resistant to the extraction by 1% Triton X-100 and 0.3 M KI were thought to represent IFs. Bershadsky et al (1978) reported that the treatment of cultured cells with 1% Triton X-100 resulted in the extraction of about 80% cellular protein, and according to Blikstad et al (1978), high proportion of the released actin is likely to be of an unpolymerized form. So it seems unlikely that treatment with 1% Triton X-100 at 3TC in the presence of 4 M glycerol and 0.1 mM EDTA affects the structure of the hepatocyte cytoskeletons.…”

Section: Electron Microscopy Of Epoxy-embedded Sectionsmentioning

confidence: 99%

Morphologic study of cytoskeletal systems of mouse hepatocytes using polyethylene glycol-embedding method.

Ishii¹,

Suzuki²,

Ohta³

et al. 1984

Tohoku J. Exp. Med.

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SM mouse livers extracted by immersion in 1% Triton X-100, or in 1% Triton X-100 followed by 0.3 M KI were studied electron microscopically using the polyethylene glycolembedding method. After extraction with 1% Triton X-100, almost all the structural components of hepatocytes remained intact and cytoplasmic filaments could be seen three-dimensionally by using stereopairs of micrographs. It was difficult, however, to discriminate microfilaments, intermediate-sized filaments and microtubules from one anoter in these specimes. By immersion in 1% Triton X-100 followed by 0.3 M KI, hepatocytes were extracted remaining only plasma membranes, vesicles and filaments. These filaments were approximately 10 nm in diameter, that is intermediate in size. They were branched and were connected with plasma membranes, especilly at desmosomes. The combination method of immersion extraction and PEG-embedding seems to be suitable for the electron microscopic observaton of the cytoskeleton of cells in situ. mouse hepatocytes; polyethylene glycol; cytoskeleton; immersion extraction; stereo electron microscopy

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“…For indirect-immunofluorescence staining, cultures were extracted with buffered 1% Triton X-100 and fixed with formaldehyde. Details of the procedures were described earlier (13,14). The following first-step antibodies were used: rabbit antibodies to actin, Abbreviations: PEG, polyethylene glycol.…”

Section: Introductionmentioning

confidence: 99%

“…§1734 solely to indicate this fact. tubulin, and fibronectin, which were described earlier (13)(14)(15), and S47D9 monoclonal antibody, reacting with vimentin and prekeratin (16 Fig. 1 i, j, and k. Focal contacts were visualized by a novel antibody-exclusion method, giving better contrast than usual interferencereflection microscopy (17).…”

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confidence: 99%

Multinucleation-induced improvement of the spreading of transformed cells on the substratum.

Lyass¹,

Bershadsky²,

Gelfand³

et al. 1984

Proc. Natl. Acad. Sci. U.S.A.

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Multinucleation of various cultured cells was produced by polyethylene glycol-induced fusion or by cytochalasin-induced block of mitosis. It was found that multinucleation induced by both methods considerably improved deficient spreading of all the tested transformed fibroblastic lines; average substratum area occupied by one cell and divided per number of nuclei was 2.0-2. (12). The mouse embryo fibroblasts and L-cell derivatives were maintained in the mixture of Eagle's basal medium and 0.5% lactalbumin hydrolysate, 1:1 (vol/vol), with 10% bovine serum. All other cell types were cultivated as described in papers cited above. Cell culture media and foetal bovine serum were from Flow Laboratories. Cycloheximide (Actidione) and cytochalasin B were from Serva (Heidelberg) and Colcemid (demecolcine) was from Sigma. To measure protein synthesis, the cultures were labeled by the addition of 0.5 ,Ci/ml (1 Ci = 3.7x 1010 Bq) of [ 4C]alanine in Earle's solution for 30 min at 37°C and then were washed consecutively by saline and 5% trichloroacetic acid for 5 min. Acid-insoluble radioactivity was measured by liquid scintillation counting.Production of Multinucleated Cells. Two methods were used.Polethylene glycol fusion. The cells washed with serumfree medium were incubated for 70 sec in 50% solutions of polyethylene glycol (PEG) (Mr, 1000; Loba Chemia, Wien, Austria) at 37°C. Then PEG was washed off by the medium, cells were incubated for 2-3 hr at 37°C and seeded on the glass coverslips at a density of about 104 cells per cm2; after 24 hr of cultivation, the cells were fixed and used for examination.Cytochalasin-induced block of cytokinesis. Cytochalasin B (2.5 ,ug/ml) was added to the culture grown on coverslips. The cells were grown with cytochalasin B for 3-5 days until numerous multinucleated cells were formed and then were transferred into the medium without cytochalasin B, incubated for 24 hr, fixed, and used for examination.Methods of Examination. For indirect-immunofluorescence staining, cultures were extracted with buffered 1% Triton X-100 and fixed with formaldehyde. Details of the procedures were described earlier (13,14). The following first-step antibodies were used: rabbit antibodies to actin, Abbreviations: PEG, polyethylene glycol. tTo whom reprint requests should be addressed. 3098The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

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Microtubules in mouse embryo fibro blasts extracted with Triton X-100

Cited by 61 publications

References 12 publications

Factors Involved in Interferon-induced or Cholera Toxin-induced Steroidogenesis in Y-1 Mouse Adrenal Tumour Cells

Factors Involved in Interferon-induced or Cholera Toxin-induced Steroidogenesis in Y-1 Mouse Adrenal Tumour Cells

Morphologic study of cytoskeletal systems of mouse hepatocytes using polyethylene glycol-embedding method.

Multinucleation-induced improvement of the spreading of transformed cells on the substratum.

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