The automated micro-axial-tomography system allows cells which have been attached to a glass fibre to be viewed from different angles. To immobilize the cells, the fibre has to be coated with a polymer offering a high positive charge density which acts as a kind of adhesive for cells.
The micro-axial-tomograph turns the glass fibre using a very small electric stepper motor (arrow). The footprint of the tomograph matches the size of a common object slide, so that it can be used with nearly any type of microscope.
To attach cells or cell nuclei onto the glass fibre, a fixture has been constructed where the fibres are firmly positioned in the centre of a glass capillary which can be filled with cell suspension using a 50-µl-pipette to attach the cells or cell nuclei.
The fibre is embedded in a medium with a suitable refraction index e.g. glycerol. It is kept precisely aligned and straightened by two V-shaped bearings one at each end.
FundamentalsThe micro-axial-tomograph allows cells to be viewed from different angles by rotating the fibre under a far-field light microscope.
Thereby one can clearly observe for example whether two Fluorescence-In-Situ-Hybridisation labelled genome regions lie above each other accidentally and thus appear as one single spot or whether they are located at a certain distance from each other.
The 3-D spatial resolution in far-field-microscopes is anisotropic. Therefore, the axial resolution is 2-3 times less, compared to the lateral resolution, even in a confocal laser scanning microscope.
After 3D-image reconstruction from at least three angles of view within a range of 120°, a considerable improvement of the 3D-resolution can be obtained so that the true distances between fluorescent labelled objects can be measured more precisely.