Scanning microscopy. Supplement最新文献_第9页

Preparation of thin, fine-grained, tantalum metal replicas for freeze-fracture electron microscopy. 制备用于冷冻断裂电子显微镜的薄而细的钽金属复制品。

Scanning microscopy. Supplement Pub Date : 1989-01-01

M J Costello, J Escaig

{"title":"Preparation of thin, fine-grained, tantalum metal replicas for freeze-fracture electron microscopy.","authors":"M J Costello, J Escaig","doi":"","DOIUrl":"","url":null,"abstract":"Two critical factors in the preparation of metal films on biological specimens are the type of metal used and the potentially damaging effects of radiant energy from the hot metal source. The excessive heating of surfaces is a major limitation to the replication of heat-sensitive aqueous specimens with refractory metals such as tungsten and tantalum, although these metals are known to form smaller grains and thinner films than the more commonly used platinum/carbon deposited under similar conditions. We describe here an electron gun designed for the evaporation of pure tantalum; surface heating is reduced through intermittent deposition controlled by varying the open/closed intervals of a fast shutter that operates in ultra-high vacuum. The effectiveness of the shutter was evaluated with a thin thermocouple in place of the specimen. The composition of the replicas was determined by x-ray microanalysis and by direct observation of the initial melting and subsequent evaporation of the tantalum bead supported on a tungsten rod that remained unchanged during the evaporation. The quality of the tantalum replicas was demonstrated with freeze-fracture replicas of reconstituted proteoliposomes and native membrane vesicles. With shutter intervals of 0.5 sec open and 1.0 sec closed, the surface heating was reduced enough to prevent unintentional etching and to preserve small pits complementary to protein particles in hydrophobic membrane surfaces and in ice.","PeriodicalId":77379,"journal":{"name":"Scanning microscopy. Supplement","volume":"3 ","pages":"189-99; discussion 199-200"},"PeriodicalIF":0.0,"publicationDate":"1989-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"13835806","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Quantitative morphology of the zinc-iodide-osmium (ZIO) stained synaptic vesicles. 碘化锌-锇(ZIO)染色突触囊泡的定量形态学。

Scanning microscopy. Supplement Pub Date : 1989-01-01

C Bertoni-Freddari, P Fattoretti, T Casoli, F Masera, W Meier-Ruge, J Ulrich

{"title":"Quantitative morphology of the zinc-iodide-osmium (ZIO) stained synaptic vesicles.","authors":"C Bertoni-Freddari, P Fattoretti, T Casoli, F Masera, W Meier-Ruge, J Ulrich","doi":"","DOIUrl":"","url":null,"abstract":"A computer assisted morphometric method has been elaborated to quantify synaptic vesicles evidenced by means of the Zinc-Iodide-Osmium (ZIO) staining procedure in nerve endings of a very discrete area of the cerebellar granular layer: the glomerulus. The following parameters were calculated directly on electron microscopic negatives of 4.67 microns 2 of surface terminal area: number of vesicles per unit area (Na), and per unit volume (Nv), volume density (Vv), average diameter (d) and average volume of the single vesicle (V). Ultrastructural changes taking place at nerve endings also cover synaptic vesicles, thus quantitative studies regarding vesicle population at synaptic regions can be correlated to functional changes occurring in the process of chemical transmission and reflect the plasticity of synaptic junctional zones. Although this histochemical staining method generally is referred to as unspecific, after comparing our data with the available literature reports, we propose that ZIO-positive vesicles could have a physiological significance. These ZIO-positive organelles could take part in the intraterminal homeostatic control of Ca++ ions.","PeriodicalId":77379,"journal":{"name":"Scanning microscopy. Supplement","volume":"3 ","pages":"101-6; discussion 107"},"PeriodicalIF":0.0,"publicationDate":"1989-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"13628560","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Cryotechniques in macromolecular research (a comparative study). 低温技术在大分子研究中的应用(比较研究)。

Scanning microscopy. Supplement Pub Date : 1989-01-01

M V Nermut, P Eason

{"title":"Cryotechniques in macromolecular research (a comparative study).","authors":"M V Nermut, P Eason","doi":"","DOIUrl":"","url":null,"abstract":"There is no single method which would provide an unambiguous image of all types of biological macromolecules. The choice of method depends largely on the size and properties of the macromolecule. Obviously small molecules are best visualized by negative staining, the problems appear with negative staining of larger structures. Here, the uncertainty about which part of the complex is actually stained (top or bottom) makes correct interpretation difficult. Shadowing techniques have the advantage of both visualizing the surface and also delineating the whole macromolecule, but suffer from lower resolution due to the graininess of the metal. However, they are superior to negative staining for the visualization of thin linear macromolecules. The next series of problems includes the interaction of macromolecules with supporting films, glass coverslips or mica, which can be hydrophobic, hydrophilic or charged and these properties can influence the orientation of the molecules. Surface tension forces during air-drying must also be considered. We have used a variety of preparative techniques in our studies of biological macromolecules: (a) negative staining; (b) air-drying from ethanol; (c) glycerol-spraying; (d) adsorption freeze-drying; (e) monolayer freeze-etching. These methods have been tested on small viruses, water soluble proteins (ribosomes, F-actin, microtubules) and transmembrane proteins requiring the presence of detergents (sarcoplasmic reticulum ATPase, fibronectin receptor). We find that freeze-drying is the most reliable and easy method for molecules that withstand distilled water; freeze-etching can be successfully applied to transmembrane proteins (even in the presence of detergents or salt); the glycerol-spray technique provides an excellent alternative to the cryotechniques in particular for studies of single linear molecules.","PeriodicalId":77379,"journal":{"name":"Scanning microscopy. Supplement","volume":"3 ","pages":"213-24; discussion 224-5"},"PeriodicalIF":0.0,"publicationDate":"1989-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"13835808","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Cryoultramicrotomy of ultra-rapidly frozen specimens. 超高速冷冻标本的冷冻切片。

Scanning microscopy. Supplement Pub Date : 1989-01-01

R L Ornberg

引用次数: 0

The organization and substructure of chromatin fibres in the interphase nucleus as studied by scanning electron microscopy. 扫描电镜研究间期细胞核染色质纤维的组织和亚结构。

Scanning microscopy. Supplement Pub Date : 1989-01-01

T D Allen

{"title":"The organization and substructure of chromatin fibres in the interphase nucleus as studied by scanning electron microscopy.","authors":"T D Allen","doi":"","DOIUrl":"","url":null,"abstract":"The high packaging ratio of DNA in both interphase nuclei and metaphase chromosomes presents great difficulties to our understanding of the three dimensional organisation of processes such as DNA replication and transcription in the nucleus. Although the higher order structure of DNA, in terms of the way it is organised into the unit fibre of chromatin has received much attention over the last decade, the highest levels of packaging of chromatin in both nuclei and chromosomes have hardly begun to be elucidated. Much of the difficulty in investigating fibre organisation with conventional methods is the inherent two dimensional nature of sectioned or spread material in the transmission microscope. Three dimensional imaging from the SEM has, until recently, been limited by the available resolution. Our own previous studies of chromosome structure have shown that a combination of 'in lens' imaging combined with the high signal generation imparted by osmium impregnation have been adequate to routinely visualise chromatin fibre organisation in metaphase chromosomes, and the changes that occur as a result of a variety of banding techniques. More recent experiments using the same techniques on interphase nuclei extracted from a variety of tissue culture cells have indicated that Scanning electron microscopy of nuclei is a potentially useful technique for studying chromatin organisation, which may be made more accessible by a variety of biochemical extraction methods.","PeriodicalId":77379,"journal":{"name":"Scanning microscopy. Supplement","volume":"3 ","pages":"77-85; discussion 85-6"},"PeriodicalIF":0.0,"publicationDate":"1989-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"13703072","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Problems associated with the preparation of whole mounts of cytoskeletons for high resolution electron microscopy. 高分辨率电子显微镜用细胞骨架全支架制备的相关问题。

Scanning microscopy. Supplement Pub Date : 1989-01-01

P B Bell, M Lindroth, B A Fredriksson, X D Liu

{"title":"Problems associated with the preparation of whole mounts of cytoskeletons for high resolution electron microscopy.","authors":"P B Bell, M Lindroth, B A Fredriksson, X D Liu","doi":"","DOIUrl":"","url":null,"abstract":"The resolution currently available in both transmission and scanning electron microscopes is theoretically adequate to visualize the organization of the cytoskeleton at the supramolecular and macromolecular levels. However, achieving this resolution in practice requires that the methods used to prepare the specimens both preserve the structures of interest and render them visible for observation in the microscope without obscuring or altering them. In this paper we discuss our own and others efforts to develop methods to overcome several problems associated with preparing whole mounts of cytoskeletons for observation by electron microscopy. These problems include: controlling the degree to which cellular components are extracted; the effects of osmium tetroxide on the cytoskeleton; controlling and recognizing shrinkage and drying artifacts; the choice of a method of visualization; deposition of grain-free ultrathin films of metal; and interpreting the results. The standard procedure which we currently use consists of the following steps: growing cells on carbon-stabilized Formvar-coated gold electron microscope grids; extracting in 0.5% Triton X-100 detergent in a microtubule stabilizing buffer; postfixing in 2.5% glutaraldehyde in stabilizing buffer; freeze-drying; magnetron sputter-coating with 1.5 nm of tungsten; and observation by TEM, SEM, or STEM. Cytoskeletons prepared in this manner contain over 100 polypeptides and are composed of a complex three dimensional meshwork of clean, uniform filaments, the smallest of which are 7 nm in diameter. A structure resembling the microtrabecular lattice is present only if the cells are prefixed with a relatively long bifunctional protein crosslinking reagent prior to extraction with detergent.","PeriodicalId":77379,"journal":{"name":"Scanning microscopy. Supplement","volume":"3 ","pages":"117-34; discussion 134-5"},"PeriodicalIF":0.0,"publicationDate":"1989-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"13835178","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Probe size and bound label conformation in colloidal gold-ligand labels and gold-immunolabels. 胶体金配体标记和金免疫标记中的探针大小和结合标记构象。

Scanning microscopy. Supplement Pub Date : 1989-01-01

S R Simmons, R M Albrecht

引用次数: 0

The contracting muscle: a challenge for freeze-substitution and low temperature embedding. 收缩肌:冷冻替代和低温包埋的挑战。

Scanning microscopy. Supplement Pub Date : 1989-01-01

L Edelmann

{"title":"The contracting muscle: a challenge for freeze-substitution and low temperature embedding.","authors":"L Edelmann","doi":"","DOIUrl":"","url":null,"abstract":"Frog sartorius and semitendinosus muscles are quick-frozen either in the resting state or during contraction by means of a LN2 cooled falling copper block. The frozen specimens are freeze-substituted (acetone + OsO4 + uranyl acetate) in a REICHERT JUNG CS auto and either embedded in Spurr's resin and polymerised at a high temperature (60 degrees C) or embedded and polymerised in the Lowicryls K4M, K11M or HM23 at low temperatures (below -30 degrees C). Excellent morphological results are obtained when freeze-substitution, embedding and polymerisation are all carried out below -50 degrees C. Muscles in which a major portion of cellular K+ ions has been replaced by electron dense Cs+ or Tl+ ions are also cryofixed at rest or during contraction, freeze-substituted in pure acetone for 1 week at -80 degrees C and polymerised in K11M at -60 degrees C. A characteristic uneven distribution of the electron dense ions--known from earlier published control experiments--can be observed in sections of resting muscles. Electrically stimulated muscles show ion redistribution. It is concluded that freeze-substitution and low temperature embedding of quick-frozen contracting muscle may be used to investigate changes of ultrastructure, redistribution of cellular water and intracellular movements of mobile ions during muscle contraction.","PeriodicalId":77379,"journal":{"name":"Scanning microscopy. Supplement","volume":"3 ","pages":"241-51; discussion 251-2"},"PeriodicalIF":0.0,"publicationDate":"1989-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"13760326","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

High pressure freezing comes of age. 高压冷冻是成熟的。

Scanning microscopy. Supplement Pub Date : 1989-01-01

D Studer, M Michel, M Müller

引用次数: 0

Characterization of chromatin structure by image analysis--a method for the assessment of changes in chromatin organization. 通过图像分析表征染色质结构——一种评估染色质组织变化的方法。

Scanning microscopy. Supplement Pub Date : 1989-01-01

H E Danielsen, G Farrants, A Reith

引用次数: 0