Abstract
The possibility to quantitatively study specific molecular/cellular features of complete human organs with preserved spatial 3D context would have widespread implications for pre-clinical and clinical medicine. Whereas optical 3D imaging approaches have experienced a formidable revolution, they have remained limited due to current incapacities in obtaining specific labelling within large tissue volumes. We present a simple approach enabling reconstruction of antibody labeled cells within entire human organs with preserved organ context. We demonstrate the utility of the approach by providing volumetric data and 3D distribution of hundreds of thousands of islets of Langerhans within the human pancreas. By assessments of pancreata from non-diabetic and type 2 diabetic individuals, we display previously unrecognized features of the human islet mass distribution and pathology. As such, this method may contribute not only in unraveling new information of the pancreatic anatomy/pathophysiology, but it may be translated to essentially any antibody marker or organ system.
Highlights
A range of mesoscopic imaging approaches are currently available for optical deep tissue imaging of biological samples, including optical projection tomography (OPT)[1] and light-sheet fluorescence microscopy (LSFM)[2,3,4]
OPT was used as the primary technology in this study, the implemented tissue processing protocols are fully compatible with LSFM, applied here to obtain complementary highresolution assessments
By computer-assisted clustering of islets ≤ 300 μm from each other in 3D space, we identified both high islet density regions (HIDR), defined as consisting of >100 islets, and low islet density regions (LIDR) Notably, HIDRs were consistently observed in the periphery of the gland in pancreata from both ND (n = 5) and type 2 diabetic (T2D) donors (n = 2) (Fig. 3a–c, Supplementary Movie 5)
Summary
A range of mesoscopic imaging approaches are currently available for optical deep tissue imaging of biological samples, including optical projection tomography (OPT)[1] and light-sheet fluorescence microscopy (LSFM)[2,3,4] (for comparative review see Liu et al.[5]) These technologies can provide micrometer resolution 3D renderings of optically cleared tissues on the mm-cm scale. The technique can be performed by presently available mesoscopic imaging devices and builds on the stitching of antibody labelled cm3-sized tissue blocks back into Solving such quantitative issues would allow for regressive indications on the endocrine organization and the relation to β-cell function of the organ. Fixed pancreas cast in agarose and cut in cm3 - sized cuboids d e Ab labelled Endogenous Merge islets Autofluorescence
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