The cytoplasmic filament system in critical point-dried whole mounts and plastic-embedded sections.

Ris, Hans

doi:10.1083/jcb.100.5.1474

Cited by 163 publications

(114 citation statements)

References 11 publications

(13 reference statements)

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“…Filaments assembled in vitro were grown on 22×22 mm ethanolwashed glass coverslips and fixed for 30 minutes in 1% glutaraldehyde, dehydrated in ethanol and critical-point dried as described previously (Ris, 1985). After sputter-coating with gold palladium, samples were imaged in a JEOL 840 scanning electron microscope operated at 20 kV.…”

Section: Electron Microscopymentioning

confidence: 99%

A 48 kDa integral membrane phosphoprotein orchestrates the cytoskeletal dynamics that generate amoeboid cell motility in Ascaris sperm

LeClaire

Stewart

Roberts

2003

Journal of Cell Science

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Protrusion of the lamellipod in the crawling sperm of Ascaris is tightly coupled to the localized vectorial assembly and bundling of the major sperm protein cytoskeleton. In cell-free extracts of sperm, vesicles derived from the leading edge membrane reconstitute protrusion by directing the assembly of columnar meshworks of major sperm protein filaments that push the vesicle forward as they elongate. Treatment with proteases or a tyrosine phosphatase abolished vesicle activity, suggesting the involvement of a membrane phosphoprotein. Fractionation of vesicle proteins by sequential detergent lysis, size exclusion chromatography and immunoprecipitation with antiphosphotyrosine antibody identified a 48 kDa integral membrane phosphoprotein as the only sperm membrane component required to nucleate major sperm protein polymerization under physiological conditions. Immunolabeling assays showed that this protein is distributed uniformly in the sperm plasma membrane, but that its active phosphorylated form is located only at sites of major sperm protein polymerization at the leading edge. Because this protein specifies sites of cytoskeletal assembly, we have named it major sperm protein polymerization organizing protein (MPOP). The phosphorylation of MPOP is pH sensitive and appears to require a soluble tyrosine kinase. Comparison of the activity of MPOP to that of analogous membrane proteins in actin-based systems emphasizes the importance of precise transmission of information from the membrane to the cytoskeleton in amoeboid cell motility.

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Section: Electron Microscopymentioning

confidence: 99%

A 48 kDa integral membrane phosphoprotein orchestrates the cytoskeletal dynamics that generate amoeboid cell motility in Ascaris sperm

LeClaire

Stewart

Roberts

2003

Journal of Cell Science

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“…Although important biological information can be extracted from NSOM on dry biological samples, these results are always subject to potential drying artifacts (35). We have recently shown that the performance of NSOM can be extended to measurements in liquid environments using a diving bell concept (34), and showed for the first time single molecule detection sensitivity with 90 nm spatial resolution on wet cells (32).…”

Section: High-resolution Nsom Imaging In Liquidmentioning

confidence: 99%

Near-Field Fluorescence Microscopy: An Optical Nanotool to Study Protein Organization at the Cell Membrane

García‐Parajó

Bakker

Koopman

et al. 2005

NBT

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The ability to study the structure and function of cell membranes and membrane components is fundamental to understanding cellular processes. This requires the use of methods capable of resolving structures with nanometer-scale resolution in intact or living cells. Although fluorescence microscopy has proven to be an extremely versatile tool in cell biology, its diffraction-limited resolution prevents the investigation of membrane compartmentalization at the nanometer scale. Near-field scanning optical microscopy (NSOM) is a relatively unexplored technique that combines both enhanced spatial resolution of probing microscopes and simultaneous measurement of topographic and optical signals. Because of the very small nearfield excitation volume, background fluorescence from the cytoplasm is effectively reduced, enabling the visualization of nano-scale domains on the cell membrane with single molecule detection sensitivity at physiologically relevant packing densities. In this article we discuss technological aspects concerning the implementation of NSOM for cell membrane studies and illustrate its unique advantages in terms of spatial resolution, background suppression, sensitivity, and surface specificity for the study of protein clustering at the cell membrane. Furthermore, we demonstrate reliable operation under physiological conditions, without compromising resolution or sensitivity, opening the road toward truly live cell imaging with unprecedented detail and accuracy.

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“…The CPD method using dry ice has an inherent disadvantage in the unchangeability of an intermediate fluid of isoamyl acetate. If our CPD was performed by the ideal method by Ris (1985), we believe there would be no differences between the two drying methods.…”

Section: Complementary Observation Of the Fractured Golgi Apparatusmentioning

confidence: 99%

“…The present evaluation using the complementary SEM confirmed that BFD preserved ultrastructures better than CPD. Ris (1985) has reported that a complete replacement of the intermediate solvent with absolutely dehydrated liquid carbon dioxide is required for attaining desirable CPD. The CPD method using dry ice has an inherent disadvantage in the unchangeability of an intermediate fluid of isoamyl acetate.…”

Section: Complementary Observation Of the Fractured Golgi Apparatusmentioning

confidence: 99%

Complementary Scanning Electron Microscopy: Technical Notes and Applications.

Inoué

1992

Archives of Histology and Cytology

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Summary.This report introduces practical techniques and applications of complementary scanning electron microscopy (SEM). To identify the complementary structures at high magnification, we first made a montage pair of low magnification micrographs as a guide map. Consulting the map, we took complementary micrographs at high magnification. When taking a picture at high magnification, we drew the outline of the most prominent structures on a transparent plastic plate attached to the cathode-ray tube of a SEM. Then another picture of the complementary structures was taken after adjusting the complementary structures to the reverse image of the plastic plate.We performed three applications of the complementary SEM: 1) complementary observation of the epithelial underside and lamina propria of the rat urinary bladder; 2) complementary observation of the fractured Golgi apparatus; and 3) evaluation of specimen drying methods.After proper digestion of the rat urinary bladder using strong alkali, the epithelium was detached from the underlying lamina propria. On the basal side of the epithelium, the meshwork of grooves were visible. The observation of the corresponding lamina propria confirmed that the grooves were occupied by blood capillaries located in the uppermost part of the lamina propria.The complementary observation of the Golgi apparatus was useful for understanding its three-dimensional architecture. The observation on both complementary fractured surfaces of the Golgi apparatus from the mouse lacrimal gland demonstrated the continuity of the Golgi stacks. It was also effective for observing both the cis and trans side of the same Golgi stack.Complementary SEM was also useful for evaluating the specimen drying method. One fractured specimen obtained by freeze-cracking was dried by the criticalpoint drying (CPD) method, and another complementary specimen was dried by the t-butyl freeze-drying method (BFD). The precise comparison of the preservation of the endoplasmic reticulum of pancreatic acinar cells indicated that the BFD technique preserved the ultrastructures better than CPD.

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The cytoplasmic filament system in critical point-dried whole mounts and plastic-embedded sections.

Cited by 163 publications

References 11 publications

A 48 kDa integral membrane phosphoprotein orchestrates the cytoskeletal dynamics that generate amoeboid cell motility in Ascaris sperm

A 48 kDa integral membrane phosphoprotein orchestrates the cytoskeletal dynamics that generate amoeboid cell motility in Ascaris sperm

Near-Field Fluorescence Microscopy: An Optical Nanotool to Study Protein Organization at the Cell Membrane

Complementary Scanning Electron Microscopy: Technical Notes and Applications.

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