Superresolution Imaging of Clinical Formalin Fixed Paraffin Embedded Breast Cancer with Single Molecule Localization Microscopy
Superresolution Imaging of Clinical Formalin Fixed Paraffin Embedded Breast Cancer with Single Molecule Localization Microscopy
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DOI：
10.1038/srep40766
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发表时间：
2017-01-18
影响因子：
4.6
通讯作者：
Gibbs, Summer L.
中科院分区：
文献类型：
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作者：
Creech, Matthew K.;Wang, Jing;Gibbs, Summer L.
数以百万计的存档的福尔马林固定石蜡包埋（FFPE）标本包含有价值的分子洞察健康和患病状态，这些状态保持在其原生超微结构中。为了在纳米尺度上诊断和治疗组织中的疾病，病理学传统上使用电子显微镜（EM），但该平台具有显著的局限性，包括成本和繁琐的样品制备。单分子定位显微镜（SMLM）的发明在光学上克服了光的衍射极限，在纳米尺度上分辨荧光标记分子，导致许多令人兴奋的生物学发现。然而，SMLM在保存组织中的应用受到限制。通过对FFPE切片在组织学厚度上进行加工的免疫荧光工作流程的调整，现在可以使用SMLM在纳米尺度上可视化细胞结构，包括个体线粒体，核层中的波动，以及人类乳腺癌标本中膜突起上的HER2受体。利用散光成像，这些结构也可以在三维空间中分辨到类似800纳米的深度。这些结果证明了SMLM在有效地揭示存档临床样本的超微结构信息方面的实用性，这可能为组织的生理病理提供分子见解，以协助使用传统的样品制备方法进行疾病诊断和治疗。
Millions of archived formalin-fixed, paraffin-embedded (FFPE) specimens contain valuable molecular insight into healthy and diseased states persevered in their native ultrastructure. To diagnose and treat diseases in tissue on the nanoscopic scale, pathology traditionally employs electron microscopy (EM), but this platform has significant limitations including cost and painstaking sample preparation. The invention of single molecule localization microscopy (SMLM) optically overcame the diffraction limit of light to resolve fluorescently labeled molecules on the nanoscale, leading to many exciting biological discoveries. However, applications of SMLM in preserved tissues has been limited. Through adaptation of the immunofluorescence workflow on FFPE sections milled at histological thickness, cellular architecture can now be visualized on the nanoscale using SMLM including individual mitochondria, undulations in the nuclear lamina, and the HER2 receptor on membrane protrusions in human breast cancer specimens. Using astigmatism imaging, these structures can also be resolved in three dimensions to a depth of similar to 800 nm. These results demonstrate the utility of SMLM in efficiently uncovering ultrastructural information of archived clinical samples, which may offer molecular insights into the physiopathology of tissues to assist in disease diagnosis and treatment using conventional sample preparation methods.