Kurzfassung

How does one visualize volume objects (1 - 20 mm) like organs or mouse embryos for biomedical applications? Several strategies like the traditional histological way (mechanical sectioning, staining and imaging of single slices, 3D image reconstruction), Micro-computed tomography (Micro-CT), and optical clearing techniques in combination with Optical Projection Tomography (OPT) or light sheet microscopy/ultramicroscopy lead to 3D volume information. These techniques differ widely in their...How does one visualize volume objects (1 - 20 mm) like organs or mouse embryos for biomedical applications? Several strategies like the traditional histological way (mechanical sectioning, staining and imaging of single slices, 3D image reconstruction), Micro-computed tomography (Micro-CT), and optical clearing techniques in combination with Optical Projection Tomography (OPT) or light sheet microscopy/ultramicroscopy lead to 3D volume information. These techniques differ widely in their demands on sample preparation, technical equipment, imaging acquisition, processing speed and leading to complementary results.The histological approach first slices the object into thin sections. Although destructive, the method is unmatched in terms of specificity; numerous well-established staining techniques target special tissue types or molecules (proteins, lipids, nucleic acids), but the imaging and reconstruction of 3D information is tedious. Making the whole object transparent by optical clearing reduces the scattering of light and opens up the way for light microscope techniques such as light-sheet/ultramicroscopy or OPT. Although the clearing techniques are time consuming (from a few days to weeks), some of them are compatible with fluorescent staining, either genetically encoded or by immunofluorescence, leading also to a high specificity. Micro-CT based on the attenuation of X-rays is routinely used for bone visualization, but soft tissue can also be visualized e.g., through iodine staining, allowing for the discrimination of bone, organs and the vascular system by different grey levels. Although immersion staining is straightforward and inexpensive, the iodine concentration, osmolarity and immersion time has to be carefully adjusted to minimize tissue shrinkage and to optimize contrast. In collaboration with the Forschungsplattform Biomaterialien, we like to establish a protocol for the 3D morphological analysis of mouse embryos. Currently, we are optimizing the fixation and soft tissue staining technique.» weiterlesen» einklappen