Label-Free Neurosurgical Pathology with Stimulated Raman Imaging

Abstract

The goal of brain tumor surgery is to maximize tumor removal without injuring critical brain structures. Achieving this goal is challenging since it can be difficult to distinguish tumor from non-tumor tissue. While standard histopathology provides information that could assist tumor delineation, it cannot be performed iteratively during surgery as freezing, sectioning, and staining of the tissue require too much time. Stimulated Raman scattering (SRS) microscopy is a powerful label-free chemical imaging technology that enables rapid mapping of lipids and proteins within a fresh specimen. This information can be rendered into pathology-like images. Although this approach has been used to assess the density of glioma cells in murine orthotopic xenografts models and human brain tumors, tissue heterogeneity in clinical brain tumors has not yet been fully evaluated with SRS imaging. Here we profile 41 specimens resected from 12 patients with a range of brain tumors. By evaluating large-scale stimulated Raman imaging data and correlating this data with current clinical gold standard of histopathology for 4,422 fields of view, we capture many essential diagnostic hallmarks for glioma classification. Notably, in fresh tumor samples we observe additional features, not seen by conventional methods, including extensive lipid droplets within glioma cells, collagen deposition in gliosarcoma, and irregularity and disruption of myelinated fibers in areas infiltrated by oligodendroglioma cells. This data is freely available in a public resource to foster diagnostic training and to permit additional interrogation. Our work establishes the methodology and provides a significant collection of reference images for label-free neurosurgical pathology.

Raman spectroscopy: techniques and applications in the life sciences

“…Ji et al were also able to obtain multi-color SRS images of human brain tumors which matched well with H&E in detecting various tumor features [257]. Lu et al later extended this idea to whole-tissue imaging of fresh samples removed during neurosurgery with SRS in reasonable measurement times suitable for use during surgery see Figure 25(a) [258]. SRS is also compatible with other nonlinear optical microscopy techniques such as CARS, second/third harmonic generation and multiphoton fluorescence which share similar instrumentation and usually require simple changes in the detection configuration for measurement.…”

Stimulated Raman Scattering: From Bulk to Nano

“…It has been suggested and demonstrated, in principle, that SRS could be a viable tool for in situ differentiation of brain tumor tissue from healthy tissue. 154–158 In addition, the SRS imaging technique has been shown to be capable of following the uptake of kinase inhibitors into living cells. 159 SRS microscopy has also been used to characterize cholesterol and lipids in atherosclerotic plaques.…”

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

“…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. Similar to SRS, mid-IR spectroscopy can provide histological imaging based on intrinsic molecular contrast, but has limited ability to perform reflectance-mode imaging of live tissue, and therefore typically requires time-consuming dehydration and/or physical sectioning of tissue [40].…”