scholarly journals INTERPRETATIONS OF ELECTRON MICROGRAPHS OF SINGLE AND SERIAL SECTIONS

1955 ◽  
Vol 1 (4) ◽  
pp. 301-314 ◽  
Author(s):  
Robley C. Williams ◽  
Frances Kallman

A method of securing serial sections for electron microscopy is described. Serial sections present certain anomalies of interpretation of a nature such that a complete and detailed three-dimensional reconstruction of the sectioned tissue cannot be made. These anomalies are discussed, as well as those which have been encountered in the interpretation of single sections. Observations of the following kinds have been made in an attempt to elucidate the interpretation of single and serial sections: differing methods of mounting adjacent sections, observation of the same section by high-angle stereoscopy, and examination of sections which have been shadowed prior to and subsequent to electron microscopy. It is found that the appearance of sections is independent of the choice of side to be placed against the formvar films. Stereoscopy shows that the appearance of fine structures is strongly dependent upon the direction of the penetrating electron beam with respect to the plane of the structures. Stereoscopy, combined with shadowing, shows quantitatively that extensive sublimation of polymer occurs upon normal exposure in the electron microscope. Observation of sections shadowed prior to electron microscopy indicates that varying amounts of material are removed between sections by the action of microtomy; i.e., it is probable that the sum of the thicknesses of several serial sections is considerably less than the total thickness of material removed from the block. It is believed that this effect, combined with the effect of sublimation, aids in explaining the failure of adjacent sections to exhibit continuity in their detailed structures.

Author(s):  
Robert Glaeser ◽  
Thomas Bauer ◽  
David Grano

In transmission electron microscopy, the 3-dimensional structure of an object is usually obtained in one of two ways. For objects which can be included in one specimen, as for example with elements included in freeze- dried whole mounts and examined with a high voltage microscope, stereo pairs can be obtained which exhibit the 3-D structure of the element. For objects which can not be included in one specimen, the 3-D shape is obtained by reconstruction from serial sections. However, without stereo imagery, only detail which remains constant within the thickness of the section can be used in the reconstruction; consequently, the choice is between a low resolution reconstruction using a few thick sections and a better resolution reconstruction using many thin sections, generally a tedious chore. This paper describes an approach to 3-D reconstruction which uses stereo images of serial thick sections to reconstruct an object including detail which changes within the depth of an individual thick section.


Author(s):  
Gregory J. Czarnota

Chromatin structure at the fundamental level of the nucleosome is important in vital cellular processes. Recent biochemical and genetic analyses show that nucleosome structure and structural changes are very active participants in gene expression, facilitating or inhibiting transcription and reflecting the physiological state of the cell. Structural states and transitions for this macromolecular complex, composed of DNA wound about a heterotypic octamer of variously modified histone proteins, have been measured by physico-chemical techniques and by enzyme-accessibility and are recognized to occur with various post-translational modifications, gene activation, transformation and with ionic-environment. In spite of studies which indicate various forms of nucleosome structure, all current x-ray and neutron diffraction studies have consistently resulted in only one structure, suggestive of a static conformation. In contrast, two-dimensional electron microscopy studies and three-dimensional reconstruction techniques have yielded different structures. These fundamental differences between EM and other ultrastructural studies have created a long standing quandary, which I have addressed and resolved using spectroscopic electron microscopy and statistical analyses of nucleosome images in a study of nucleosome structure with ionic environment.


Author(s):  
Nicolas Boisset ◽  
Jean-Christophe Taveau ◽  
Jean Lamy ◽  
Terence Wagenknecht ◽  
Michael Radermacher ◽  
...  

Hemocyanin, the respiratory pigment of the scorpion Androctonus australis is composed of 24 kidney shaped subunits. A model of architecture supported by many indirect arguments has been deduced from electron microscopy (EM) and immuno-EM. To ascertain, the disposition of the subunits within the oligomer, the 24mer was submitted to three-dimensional reconstruction by the method of single-exposure random-conical tilt series.A sample of native hemocyanin, prepared with the double layer negative staining technique, was observed by transmisson electron microscopy under low-dose conditions. Six 3D-reconstructions were carried out indenpendently from top, side and 45°views. The results are composed of solid-body surface representations, and slices extracted from the reconstruction volume.The main two characters of the molecule previously reported by Van Heel and Frank, were constantly found in the solid-body surface representations. These features are the presence of two different faces called flip and flop and a rocking of the molecule around an axis passing through diagonnally opposed hexamers. Furthermore, in the solid-body surface of the top view reconstruction, the positions and orientations of the bridges connecting the half molecules were found in excellent agreement with those predicted by the model.


Reproduction ◽  
2012 ◽  
Vol 144 (3) ◽  
pp. 385-392 ◽  
Author(s):  
Unai Silván ◽  
Juan Aréchaga

Cell transplantation into the seminiferous tubules is a useful technique for the study of physiological and pathological conditions affecting the testis. However, the precise three-dimensional organization and, particularly, the complex connectivity of the seminiferous network have not yet been thoroughly characterized. To date, the technical approaches to address these issues have included manual dissection under the stereomicroscope, reconstruction of histological serial sections, and injection of contrast dyes, but all of them have yielded only partial information. Here, using an approach based on the microinjection of a self-polymerizing resin followed by chemical digestion of the surrounding soft tissues, we reveal fine details of the seminiferous tubule scaffold and its connections. These replicas of the testis seminiferous network were studied by scanning electron microscopy. The present results not only establish a morphological basis for more precise microinjection into the mouse seminiferous tubules but also enable a more profound investigation of physiological and embryological features of the testis.


2012 ◽  
pp. 881-898
Author(s):  
J.R. Bilbao-Castro ◽  
I. García ◽  
J.J. Fernández

Three-dimensional electron microscopy allows scientists to study biological specimens and to understand how they behave and interact with each other depending on their structural conformation. Electron microscopy projections of the specimens are taken from different angles and are processed to obtain a virtual three-dimensional reconstruction for further studies. Nevertheless, the whole reconstruction process, which is composed of many different subtasks from the microscope to the reconstructed volume, is not straightforward nor cheap in terms of computational costs. Different computing paradigms have been applied in order to overcome such high costs. While classic parallel computing using mainframes and clusters of workstations is usually enough for average requirements, there are some tasks which would fit better into a different computing paradigm – such as grid computing. Such tasks can be split up into a myriad of subtasks, which can then be run independently using as many computational resources as are available. This chapter explores two of these tasks present in a typical three-dimensional electron microscopy reconstruction process. In addition, important aspects like fault-tolerance are widely covered; given that the distributed nature of a grid infrastructure makes it inherently unstable and difficult to predict.


Author(s):  
J.R. Bilbao Castro ◽  
I. Garcia Fernandez ◽  
J. Fernandez

Three-dimensional electron microscopy allows scientists to study biological specimens and to understand how they behave and interact with each other depending on their structural conformation. Electron microscopy projections of the specimens are taken from different angles and are processed to obtain a virtual three-dimensional reconstruction for further studies. Nevertheless, the whole reconstruction process, which is composed of many different subtasks from the microscope to the reconstructed volume, is not straightforward nor cheap in terms of computational costs. Different computing paradigms have been applied in order to overcome such high costs. While classic parallel computing using mainframes and clusters of workstations is usually enough for average requirements, there are some tasks which would fit better into a different computing paradigm – such as grid computing. Such tasks can be split up into a myriad of subtasks, which can then be run independently using as many computational resources as are available. This chapter explores two of these tasks present in a typical three-dimensional electron microscopy reconstruction process. In addition, important aspects like fault-tolerance are widely covered; given that the distributed nature of a grid infrastructure makes it inherently unstable and difficult to predict.


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