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"Electron microscopy and stereology in cell biology "

An EMBO course held jointly with FEBS


20th - 29th June | 2006 | Oslo| Norway

 

FEBS
Supported by FEBS
Youth Travel Grant

About the Course

 

The aim of this course is to teach cutting-edge EM techniques for Cell Biologists on a high theoretical and practical level. EM is technically one of the most demanding set of approaches to learn; many aspects can only reasonably be understood by watching a specialist. Despite the vast number of technical books available, there is clearly no substitute for hands-on training.
Therefore the main emphasis in this course is put on the practical training.

Since their invention of transmission electron microscope in the 1930’s electron microscopes have been using for studying the in situ functional organization of the cell. Nowadays conventional EM gives relatively little information about the relation between a structure and its function in a cell, since it cannot identify specific structures and then ascribe specific processes to them. Routinely we can observe only a few dozens of structural components in the interior of the cell. These components cannot account for a description of the thousands of functional cellular processes taking place at any one moment. In order to gain a deeper understanding of structure-function relationships in the cell, it is necessary to implement affinity cytochemistry at the EM level. Ultrastructural immunocytochemistry represents the most definite technique to identify specific cell structures in situ with sufficient spatial resolution.

Recently, the use of these EM approaches has become less prominent. Likely reasons for this phenomenon are the increasing power of confocal and video light microscopy on the one hand, on the other hand the expense and working difficulties of EM techniques have effect also on this downtrend. However, only EM techniques can bring the information about antigen localization at the ultrastructural level, when is necessary to distinguish between the many interesting organelles whose functions are coupled (e.g. Golgi, ER). EM approaches complement light microscopy by providing data unavailable through other methods.

The course has a long tradition because it has been organized annually for about 20 years. During this period a stable teaching team has been formed who have great experience with teaching these techniques. It is composed of the world’s best specialists in processing of cells and tissues for EM in combination with immunogold labelling and quantitation via stereology.
The teachers are able to teach someone with no previous experience in EM to a level where the students can not only establish the techniques in their home labs but also teach others. Many of the students are exposed for the first time to any formal teaching in understanding three-dimensional structures from two-dimensional sections. For this one uses state-of-the-art stereology. This approach passed through a great evolution during the past 20 years, a period which has brought new methods to sample biological material efficiently and in an unbiased fashion and thus to obtain reliable estimates of surface, volume, length or number of any visible structure. A part of the course is therefore dedicated to stereological analysis, a process that allows quantification of the results. It has particular importance because it shifts the subjective evaluation of experiments to the objective level. It is still surprising how such an elegant and effective set of methods are unknown to most cell biologists, including the majority of EM specialists.

In the course the students learn the best methods for preserving, visualizing and localizing antigens on cellular structures. They are encouraged to bring their own specimens and antibodies to the course. The Tokuyasu cryo section method is very rapid in comparison with routine imunolabelling on resin sections and allows some students to achieve publishable data during the course.
The use of email these days also facilitates continued interactions between the students and the teachers after the course; this has been especially prominent during the past three years.

Another very important topic that we cover is the boundary between light microscopy and electron microscopy. For many years Heinz Schwarz has been emphasizing the power of using EM-style ultrathin sections for high resolution light microscopy immunocytochemistry. This is a poorly appreciated, albeit simple, approach that can routinely provide a significantly higher resolution than the confocal microscope