Wilson Thomas J, Toland Angus, Cayrol Romain, Vogel Hannes
Department of Neurosurgery, Stanford University, Stanford, CA, USA.
Department of Pathology, Stanford University, Stanford, CA USA.
Clin Neurol Neurosurg. 2022 Mar;214:107180. doi: 10.1016/j.clineuro.2022.107180. Epub 2022 Feb 18.
During nerve repair, an intraoperative assessment of the quality of the nerve stump is critically important for achieving a good outcome. Frozen section analysis of osmium-hematoxylin stained sections has not been adopted at many centers, including ours. This has left us with bread-loafing the nerve and visually assessing for healthy fascicles. A technique that would allow for rapid, safe, quantitative intraoperative assessment of nerve quality, including myelin quantity, would be beneficial. Stimulated Raman Scattering (SRS) microscopy is a rapid, label-free technique that images lipids well that may be uniquely suited for this purpose.
To describe our initial experience and lessons learned using SRS microscopy for evaluation of peripheral nerve tissue.
We present 6 cases during which SRS microscopy was used to evaluate peripheral nerve tissue, including standard histology and SRS microscopy images, where applicable.
Our current technique involves OCT embedding the nerve tissue and then cutting 70 µm sections on a standard cryostat. The SRS microscope slide is modified to change the buffer depth from 100 µm to 50 µm. We analyzed the gray scale composite images, merged from the CH (lipid) channel and CH (protein) channel. This technique reliably produced cross-sectional images and showed good capability for imaging myelinated axons within fascicles.
We demonstrate here an innovative approach to quantifying myelin in peripheral nerve using Stimulated Raman Scattering microscopy. This should prove useful in the care and surgical treatment of patients with peripheral nerve injuries.
在神经修复过程中，术中对神经残端质量进行评估对于取得良好疗效至关重要。包括我们中心在内的许多机构尚未采用锇苏木精染色切片的冷冻切片分析方法。这使得我们只能将神经切成面包片样并通过肉眼评估健康的神经束。一种能够在术中快速、安全且定量地评估神经质量（包括髓磷脂含量）的技术将大有裨益。受激拉曼散射（SRS）显微镜检查是一种快速、无需标记的技术，能够很好地对脂质成像，可能特别适用于此目的。
描述我们使用SRS显微镜评估周围神经组织的初步经验和教训。
我们展示了6例使用SRS显微镜评估周围神经组织的病例，包括适用时的标准组织学和SRS显微镜图像。
我们目前的技术包括将神经组织进行OCT包埋，然后在标准低温恒温器上切成70μm的切片。对SRS显微镜载玻片进行了改进，将缓冲液深度从100μm改为50μm。我们分析了从CH（脂质）通道和CH（蛋白质）通道合并的灰度合成图像。该技术可靠地生成了横截面图像，并显示出对神经束内有髓轴突成像的良好能力。
我们在此展示了一种使用受激拉曼散射显微镜定量周围神经髓磷脂的创新方法。这在周围神经损伤患者的护理和手术治疗中应会证明是有用的。
