scholarly journals A short primer on lung stereology

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Matthias Ochs ◽  
Julia Schipke

AbstractThe intention of this short primer is to raise your appetite for proper quantitative assessment of lung micro-structure. The method of choice for obtaining such data is stereology. Rooted in stochastic geometry, stereology provides simple and efficient tools to obtain quantitative three-dimensional information based on measurements on nearly two-dimensional microscopic sections. In this primer, the basic concepts of stereology and its application to the lung are introduced step by step along the workflow of a stereological study. The integration of stereology in your laboratory work will help to improve its quality. In a broader context, stereology may also be seen as a contribution to good scientific practice.

Author(s):  
H.A. Cohen ◽  
T.W. Jeng ◽  
W. Chiu

This tutorial will discuss the methodology of low dose electron diffraction and imaging of crystalline biological objects, the problems of data interpretation for two-dimensional projected density maps of glucose embedded protein crystals, the factors to be considered in combining tilt data from three-dimensional crystals, and finally, the prospects of achieving a high resolution three-dimensional density map of a biological crystal. This methodology will be illustrated using two proteins under investigation in our laboratory, the T4 DNA helix destabilizing protein gp32*I and the crotoxin complex crystal.


Author(s):  
B. Ralph ◽  
A.R. Jones

In all fields of microscopy there is an increasing interest in the quantification of microstructure. This interest may stem from a desire to establish quality control parameters or may have a more fundamental requirement involving the derivation of parameters which partially or completely define the three dimensional nature of the microstructure. This latter categorey of study may arise from an interest in the evolution of microstructure or from a desire to generate detailed property/microstructure relationships. In the more fundamental studies some convolution of two-dimensional data into the third dimension (stereological analysis) will be necessary.In some cases the two-dimensional data may be acquired relatively easily without recourse to automatic data collection and further, it may prove possible to perform the data reduction and analysis relatively easily. In such cases the only recourse to machines may well be in establishing the statistical confidence of the resultant data. Such relatively straightforward studies tend to result from acquiring data on the whole assemblage of features making up the microstructure. In this field data mode, when parameters such as phase volume fraction, mean size etc. are sought, the main case for resorting to automation is in order to perform repetitive analyses since each analysis is relatively easily performed.


Author(s):  
Yu Liu

The image obtained in a transmission electron microscope is the two-dimensional projection of a three-dimensional (3D) object. The 3D reconstruction of the object can be calculated from a series of projections by back-projection, but this algorithm assumes that the image is linearly related to a line integral of the object function. However, there are two kinds of contrast in electron microscopy, scattering and phase contrast, of which only the latter is linear with the optical density (OD) in the micrograph. Therefore the OD can be used as a measure of the projection only for thin specimens where phase contrast dominates the image. For thick specimens, where scattering contrast predominates, an exponential absorption law holds, and a logarithm of OD must be used. However, for large thicknesses, the simple exponential law might break down due to multiple and inelastic scattering.


Author(s):  
D. E. Johnson

Increased specimen penetration; the principle advantage of high voltage microscopy, is accompanied by an increased need to utilize information on three dimensional specimen structure available in the form of two dimensional projections (i.e. micrographs). We are engaged in a program to develop methods which allow the maximum use of information contained in a through tilt series of micrographs to determine three dimensional speciman structure.In general, we are dealing with structures lacking in symmetry and with projections available from only a limited span of angles (±60°). For these reasons, we must make maximum use of any prior information available about the specimen. To do this in the most efficient manner, we have concentrated on iterative, real space methods rather than Fourier methods of reconstruction. The particular iterative algorithm we have developed is given in detail in ref. 3. A block diagram of the complete reconstruction system is shown in fig. 1.


Author(s):  
J. Silcox ◽  
R. H. Wade

Recent work has drawn attention to the possibilities that small angle electron scattering offers as a source of information about the micro-structure of vacuum condensed films. In particular, this serves as a good detector of discontinuities within the films. A review of a kinematical theory describing the small angle scattering from a thin film composed of discrete particles packed close together will be presented. Such a model could be represented by a set of cylinders packed side by side in a two dimensional fluid-like array, the axis of the cylinders being normal to the film and the length of the cylinders becoming the thickness of the film. The Fourier transform of such an array can be regarded as a ring structure around the central beam in the plane of the film with the usual thickness transform in a direction normal to the film. The intensity profile across the ring structure is related to the radial distribution function of the spacing between cylinders.


Author(s):  
A.M. Jones ◽  
A. Max Fiskin

If the tilt of a specimen can be varied either by the strategy of observing identical particles orientated randomly or by use of a eucentric goniometer stage, three dimensional reconstruction procedures are available (l). If the specimens, such as small protein aggregates, lack periodicity, direct space methods compete favorably in ease of implementation with reconstruction by the Fourier (transform) space approach (2). Regardless of method, reconstruction is possible because useful specimen thicknesses are always much less than the depth of field in an electron microscope. Thus electron images record the amount of stain in columns of the object normal to the recording plates. For single particles, practical considerations dictate that the specimen be tilted precisely about a single axis. In so doing a reconstructed image is achieved serially from two-dimensional sections which in turn are generated by a series of back-to-front lines of projection data.


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