期刊
JOURNAL OF HISTOCHEMISTRY & CYTOCHEMISTRY
卷 61, 期 4, 页码 263-271出版社
SAGE PUBLICATIONS LTD
DOI: 10.1369/0022155412473757
关键词
micro-CT; correlative microscopy; light microscopy; electron microscopy; adipose tissue; fat
类别
资金
- Shriners Hospitals for Children
- National Institutes of Health [P01 AR049698]
- Deutsche Forschungsgemeinschaft [SE 1115/1-2]
- [SFB829]
We present a method in which a precise region of interest within an intact organism is spatially mapped in three dimensions by non-invasive micro-computed X-ray tomography (micro-CT), then further evaluated by light microscopy (LM) and transmission electron microscopy (TEM). Tissues are prepared as if for TEM including osmium fixation, which imparts soft tissue contrast in the micro-CT due to its strong X-ray attenuation. This method may therefore be applied to embedded, archived TEM samples. Upon selection of a two-dimensional (2-D) projection from a region of interest (ROI) within the three-dimensional volume, the epoxy-embedded sample is oriented for microtomy so that the sectioning plane is aligned with the micro-CT projection. Registration is verified by overlaying LM images with 2-D micro-CT projections. Structures that are poorly resolved in the micro-CT may be evaluated at TEM resolution by observing the next serial ultrathin section, thereby accessing the same ROI by all three imaging techniques. We compare white adipose tissue within the forelimbs of mice harboring a lipid-altering mutation with their littermate controls. We demonstrate that individual osmium-stained lipid droplets as small as 15 mu m and separated by as little as 35 mu m may be discerned as separate entities in the micro-CT, validating this to be a high-resolution, non-destructive technique for evaluation of fat content.
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