Download e-book for iPad: Physics of Schottky Electron Sources: Theory and Optimum by Merijntje Bronsgeest. This publication comprehensively covers the Schottky emitter, facing its theoretical in addition to sensible features. The Evolution of the Microscope.
J Anat , Prog Neurobiol 37, Br J Radiol 73, Recommendations for straightforward and rigorous methods of counting neurons based on a computer simulation approach. Oct;20 1 , Cereb Cortex 12, Design-based stereology in neuroscience. J Neurocytol 33, Histol Histopathol 17, Neurobiol Aging 14, Trends Neurosci 22, Folia Morphol Warsz 60, In this way, all of the time spent on the confocal microscope is devoted to acquiring images.
There are a number of advantages to separating acquisition from analysis. Each component can be undertaken by personnel with different skill levels; the acquisition can be performed by a skilled microscopist who is familiar with the tissue, while the subsequent analysis can be performed by less skilled personnel, such as students, who only need to be trained on cell identification and the stereological counting rules.
By setting acquisitions to run in an unattended fashion see below , the imaging portion could be scheduled for times of low demand for the confocal microscope, such as overnight and on weekends. Permanent storage of the images and counting means that the identity of counted cells can be verified at a later time point. These images stacks could also be revisited at a later time point to explore other cell relationships.
The separation of acquisition and analysis also benefits collaborative projects, particularly those at a distance as acquisition can be performed at one site and analysis at another. Incidentally, it is possible to collect image stacks by brightfield microscopy, bringing these same benefits to any stereological study.
Implicit in this approach is the use of specialized software that controls the acquisition process discussed below and allows the interaction of a stereological probe with the image.
Although it is possible to conduct a study without specialized stereology software, this becomes very difficult to actually perform at high resolution. These challenges for performing confocal stereology can be overcome by using specialized commercial stereology available from the market leaders MBF Bioscience, Inc. Questions about the current capacity of software and interaction with particular confocal microscopes should be directed to these vendors.
Jacopo Annese, Arthur W.
Toga, in Brain Mapping: The Methods Second Edition , The principal limitation of conventional stereology is that the estimates are not linked in any way to larger images of the tissue that are under examination Glaser and Glaser, The application of 3D probes requires the use of high-power objective lenses with a low numerical aperture. This means that the field of view will be extremely small and not necessarily representative of the morphology of the whole section.
Semiautomated computer-based stereology systems have recently been designed to sample a distribution of fields systematically, while maintaining a record of the position and area of each sampled field relative to the entire section. The experimenter has control over the sampling procedure and determines the boundaries of the structure under examination. The microtome stage is equipped with an x—y step motor that is controlled by a computer.
The travel range of the motorized stage has to be chosen in order to cover the entire section. Large computer-controlled stages are commercially available. Although this equipment has been designed for industrial applications, it is the most suitable to survey whole sections of the human brain microscopically. Low-resolution digitization of whole sections provides the map for the navigation of the stereological probes.
This not only is practical in the analysis of one individual specimen, but also establishes the basis for a common histological framework for independent studies. The principles of stereology have been mainly applied to light microscopy, but the method is essentially valid for the analysis of data acquired by other imaging modalities such as confocal microscopy Peterson, , CT , and magnetic resonance imaging , Artificial edges are the demons of stereology , but fortunately there are only a few designs where it is necessary to sample across them.
It is a weird problem. It may be taken as a metaphor of the malice inborn in artificial edges that the few designs in this report require three different ways of handling artificial edges. It is evident that counting across all sections for an animal has to be performed before the ufasf can be estimated. Sampling is restricted to a band of fixed and constant arbitrary height, H sampl , which is divisible by the height of the frame and which is less than the local height H i.
Cell volume estimated by design-based stereology is considered as a state-of-the-art, assumption-free, and unbiased procedure for estimating neuron size, when compared to 2D or 3D particle shape-dependent methods Ladd et al.