Light-sheet microscopy for slide-free non-destructive pathology of large clinical specimens

Abstract

For the 1.7 million patients per year in the U.S. who receive a new cancer diagnosis, treatment decisions are largely made after a histopathology exam. Unfortunately, the gold standard of slide-based microscopic pathology suffers from high inter-observer variability and limited prognostic value due to sampling limitations and the inability to visualize tissue structures and molecular targets in their native 3D context. Here, we show that an open-top light-sheet microscope optimized for non-destructive slide-free pathology of clinical specimens enables the rapid imaging of intact tissues at high resolution over large 2D and 3D fields of view, with the same level of detail as traditional pathology. We demonstrate the utility of this technology for various applications: wide-area surface microscopy to triage surgical specimens (with ~200 μm surface irregularities), rapid intraoperative assessment of tumour-margin surfaces (12.5 sec/cm2), and volumetric assessment of optically cleared core–needle biopsies (1 mm in diameter, 2 cm in length). Light-sheet microscopy can be a versatile tool for both rapid surface microscopy and deep volumetric microscopy of human specimens.

Three‐dimensional reconstruction of prostate cancer architecture with serial immunohistochemical sections: hallmarks of tumour growth, tumour compartmentalisation, and implications for grading and heterogeneity

“…and Van Royen et al ., and that, according to Boag et al ., led to the failure of reconstructions in low‐grade prostate cancer. There are also some other prospective microscopy methods, which have still not been extensively tested for 3D reconstructions of prostate cancer architecture; these should be analysed in further studies …”

“…There are also some other prospective microscopy methods, which have still not been extensively tested for 3D reconstructions of prostate cancer architecture; these should be analysed in further studies. [11][12][13] However, the subtle details of the architecture (e.g. necessary for grading) are very well depicted with reconstructions involving a minimal depth of tissue.…”

Multiscale nonlinear microscopy and widefield white light imaging enables rapid histological imaging of surgical specimen margins

“…Because physical sectioning accounts for the vast majority of processing delay for both FFPE and FSA histology, optical sectioning can dramatically reduce processing times and enable intraoperative histological examination in scenarios where FFPE and FSA histology are too time consuming. Various methods have been proposed for imaging breast, prostate and other surgical margins without physical sectioning, including optical coherence tomography (OCT) [18][19][20][21][22], reflectance confocal microscopy (RCM) [23,24], confocal fluorescence microscopy (CFM) [25][26][27][28][29], structured illumination microscopy (SIM) [30], light sheet microscopy [31], microscopy with ultraviolet surface excitation (MUSE) [32,33], and nonlinear microscopy (NLM) [34][35][36][37][38]. Stimulated Raman scattering (SRS) has also been demonstrated for surgical histology [39], but typically has appreciably lower imaging speed or signal to noise ratio when performed without physical sectioning in reflectance mode.…”

Widefield light sheet microscopy using an Airy beam combined with deep-learning super-resolution

“…Widefield optical imaging at depth has opened up new vistas for neuroscience and developmental biology [1,2,3], and is expanding its remit into histopathology [4]. There, the capacity to image across length scales is an important pursuit, seeking to maximise the field of view, whilst capturing the fine details needed to discern cellular-scale morphology in three dimensions [5].…”