AIMS: CLARITY is a novel technique which enables three-dimensional visualization of immunostained tissue for the study of circuitry and spatial interactions between cells and molecules in the brain. In this study, we aimed to compare methodological differences in the application of CLARITY between rodent and large human post mortem brain samples. In addition, we aimed to investigate if this technique could be used to visualize Lewy pathology in a post mortem Parkinson's brain.
METHODS: Rodent and human brain samples were clarified and immunostained using the passive version of the CLARITY technique. Samples were then immersed in different refractive index matching media before mounting and visualizing under a confocal microscope.
RESULTS: We found that tissue clearing speed using passive CLARITY differs according to species (human vs. rodents), brain region and degree of fixation (fresh vs. formalin-fixed tissues). Furthermore, there were advantages to using specific refractive index matching media. We have applied this technique and have successfully visualized Lewy body inclusions in three dimensions within the nucleus basalis of Meynert, and the spatial relationship between monoaminergic fibres and Lewy pathologies among nigrostriatal fibres in the midbrain without the need for physical serial sectioning of brain tissue.
CONCLUSIONS: The effective use of CLARITY on large samples of human tissue opens up many potential avenues for detailed pathological and morphological studies.
|Number of pages||15|
|Journal||Neuropathology and Applied Neurobiology|
|Publication status||Published - 1 Oct 2016|
- Histocytological Preparation Techniques/methods
- Imaging, Three-Dimensional/methods
- Lewy Bodies/pathology
- Mice, Inbred C57BL
- Parkinson Disease/pathology
- Rats, Sprague-Dawley