摘要:

AbstractA guillotine‐type quick freezing device and a bio‐hazard containment box have been designed, constructed, and used to prepare vitrifiedhydrated specimens of viruses in their native environment. Special design considerations include the preservation of the specimen in its natural state in vitrified ice, prevention of virus aerosols escape, and control of the potentially explosive air‐coolant vapor mix

ISSN:0741-0581    

DOI:10.1002/jemt.1060080402

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractWater can be a substantial proportion of the residual gas in modern electron microscopes even when frozen hydrated specimens are not used. During measurements of the mass thickness of thin collodion film specimens at low temperatures, it was found that a volatile surface layer (condensed water) modified the apparent rate of mass loss induced by radiation exposure. Mass loss can be enhanced by the presence of water (specimen “etching”), or mass loss can be masked by the dynamic adsorption of water to the specimen surface. The microscope or the grid can be a secondary source of the water; even with cold anticontaminator plates in the vicinity of the specimen, water can be desorbed by x‐rays or backscattered electrons. In one typical situation, the mass loss rate appears reduced (due to water adsorption), but the ultimate damage is greater (due to etching). These results illustrate that care must be taken in interpreting mass thickness measurements made in the presence of water and that the lowest stage temperature does not necessarily produce the best observation conditions for all spec

ISSN:0741-0581    

DOI:10.1002/jemt.1060080403

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractIon milling is commonly used to prepare specimens for observation under transmission electron microscope (TEM). This technique sometimes introduces artifacts in specimens contributing to misleading interpretation of TEM results as observed in the present investigation of Al‐Li‐Cu alloys. This type of alloy, in general, contains several kinds of precipitates, namely δ′ T1, and θ′. It is found that ion milling even for a short time produces drastic changes in the precipitate characterics as compared to standard electropolishing methods of specimen preparation for TEM. Careful analysis of selected area diffraction patterns and micrographs shows that after ion milling δ′ precipitates are very irregular, whereas other precipitates coarsen and they are surrounded by misfit dislocations. In situ hot‐stage TEM experiments were performed to relate the microstructure to that observed in the ion‐milled specimen. Results and causes of ion milling effects on the microstructure are discussed in relation to standard electropolishing techniques and in situ hot

ISSN:0741-0581    

DOI:10.1002/jemt.1060080404

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractRecently, cytochemical techniques have been applied for localizing membrane components; however, transmission electron microscopy only provides two‐dimensional information about their distribution. Scanning electron microscopy, on the other hand, offers the possibility of examining the three‐dimensional architecture of biological samples. The fracture‐label cytochemical technique was combined with the backscattered electron imaging (BEI) mode of the scanning electron microscope to visualize the in vivo distribution of lectin binding sites on freeze‐fractured biological membranes in tissues and cells. Pancreatic and testicular tissues, fixed with glutaraldehyde, were freeze‐fractured and labeled withHelix pomatialectin‐gold orRicinus communisI‐gold complexes. The labeled specimens were then critical‐point dried and replicated with platinum‐carbon for routine transmission electron microscopy or with carbon alone for BEL. Lectin‐gold labeling of fractured plasma and intracellular membranes observed with BEI showed a labeling pattern similar to that seen by the replica method. However, BEI‐fracturelabel provided additional information about the distribution of the labeling with respect to three‐dimensional organization of tissues and cells. Large sample areas could be examined, making this technique particularly useful as a survey method for specimens that are either differentially labeled or composed of hetero

ISSN:0741-0581    

DOI:10.1002/jemt.1060080405

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractThe enzyme‐linked colloidal gold affinity labelling technique was tested as a method to localize cellulose on thin sections of plant cell walls and slime mold spores. Commercially available cellulase from cultures ofTrichoderma reesei, the main components being cellobiohydrolase I and II (CBH I, CBH II) and endoglucanase (EG), was linked to colloidal gold by using standard techniques and applied as a dilute, buffered suspension to thin sections. After brief exposure, e.g., 15–30 minutes, cellulose exposed on the surface of sections was labelled with the enzyme‐gold complex. Poststaining did not appear to have a deleterious effect on the labelled sections. The specificity of labelling was demonstrated by its complete inhibition when carboxymethylcellulose was incorporated in the labelling mixture, by lack of labelling of 1,4‐β‐mannans or 1,3‐β‐xylans in noncellulosic walls of marine algae, by lack of labelling of 1,4‐β‐glucans in chitin, by much lower labelling density when done at 4°C, and by lack of labelling when sections were predigested with cellulase. Labelling with the crude commercial cellulase was compared to labelling with purified CBH I‐, CBH II‐, and EG‐linked colloidal gold, and the labelling pattern was similar. This method was found useful on conventionally fixed material and required no special preparation other than the use of inert (Ni or Au) grids and 0.5% gelatin to reduce nonspecific binding of the gold complex. Labelling was similar in the several embedding resins tested: LR White, Lowicryl K4M, Epon 812, and Spurr's. The cellulase‐gold probe remained active for at least 4 weeks at 4°C and much longer when frozen at −80°C in 20% glycerol. This technique should prove useful in studies of cellulose degradation and cellulose deposition and of the interaction of cel

ISSN:0741-0581    

DOI:10.1002/jemt.1060080406

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractSome aspects of digitization of electron micrographs have been investigated. The performances of a flat‐bed, a rotating drum, and a diode array scanner have been evaluated. Estimates have been achieved for resolution, mechanical and optical stability, and optical density response. It is concluded that for routine transmission electron microscopy of, for example, negatively stained biologic specimens, a diode array scanner produces data good enough to obtain resolutions at a level normally expected. High speed is the major advantage with this type of equipment. However, for high‐resolution work it is necessary to use a conventional scanner with a relatively slow scan sp

ISSN:0741-0581    

DOI:10.1002/jemt.1060080407

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractA technique is described for preparing uniform, durable phosphor layers for viewing screens suitable for the transmission electron microscope; a settling procedure is used. The example described here is for a high‐voltage instrument, but with adjustment of the coating density, the technique should be equally suitable for screen preparation for transmission electron microscopes that operate at lower acceleration voltage

ISSN:0741-0581    

DOI:10.1002/jemt.1060080408

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractIn spite of the fact that the preembedding method is satisfactory for the ultrastructural localization of cytoskeletal proteins, there is a need for a localization method that retains the cells' ground substance, delicate filament arrangements, and membrane‐filament interactions and provides a good delineation of ultrastructural detail. Ultracryomicrotomy, a resinless sectioning method, can combine good morphology with optimal antibody labeling. Until now, however, it has not been possible to section cell monolayers parallel to their plane of growth. This is a prerequisite for the localization of proteins along segments of filaments, contained within the section thickness. We describe such a method and give a first appreciation of its potential for antibody localization studies of cytoskeletal proteins. The method consists of seeding cells on a parallel 0.75‐mm‐thick gelatin substrate that can later be cut and used as a mounting block. An adapted negative staining has yielded a very useful delineation of the well‐preserved structures within the cells, even in combination with immunogold labeling. The latter has been in its indirect version less satisfactory in dense microfilament bundles because of penetration problems, and more satisfactory on microtubules. Clearly, the penetration properties of gold probes will have to be improved before this method will become widely applicable. The availability of a sectioning method like this will provide the basis for further progress. There will be many cases which will justify the use of this relatively more difficult a

ISSN:0741-0581    

DOI:10.1002/jemt.1060080409

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractThe present review deals with the use of electron microscopy in the identification of pituitary cell types as well as the assessment of their functional state, in rat and man. Application of immunoelectron microscopy, especially immunogold techniques, utilizing multiple labeling in establishing differentiation and hormone content of cell types, is emphasized. Recent evidence of plurihormonality in various pituitary cell types indicates that the once axiomatic one cell‐one hormone theory is untenable and that the present perception of pituitary cell types and their function requires modification. Detection of hormonal and nonhormonal substances in pituitary cell types, not associated with their known endocrine function, suggests that hypophysial cells may have yet unknown roles, possibly in the realm of paracrine and autocrine regulatio

ISSN:0741-0581    

DOI:10.1002/jemt.1060080410

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractFocus in transmission electron microscopy, especially at low‐to‐moderate powers, is usually achieved by an empirical, though largely haphazard, process of defocusing the objective below what is known to be perfect focus as defined by the absence of Fresnel diffraction fringes at the edges of holes. In this article we provide a precise, empirical method of focus, whose rationale resides in the less‐than‐perfect image transfer from the image space of the objective to the negative, i.e., the modulation transfer function. In practice, “perfect focus,” as defined above, is established with a beam deflector (“wobbler”), to which underfocus is then applied routinely by reference to a table. Such “critical underfocus” values have to be calibrated, as described here, for each microscope and depend on all factors that influence contrast, that is, accelerating voltage, condenser defocus and coherence of illumination, focal length of the objective, aperture sizes, combination of intermediate and projection lenses, and the properties of the ph

ISSN:0741-0581    

DOI:10.1002/jemt.1060080411

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY