Application of direct stochastic optical reconstruction microscopy (dSTORM) to the histological analysis of human glomerular disease.
Application of direct stochastic optical reconstruction microscopy (dSTORM) to the histological analysis of human glomerular disease.
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直接随机光学重建显微镜(DSTORM)在人类肾小球疾病组织学分析中的应用
DOI：
10.1002/cjp2.217
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发表时间：
2021-09
期刊：
影响因子：
--
通讯作者：
French PM
中科院分区：
文献类型：
--
作者：
Garcia E;Lightley J;Kumar S;Kalita R;Gőrlitz F;Alexandrov Y;Cook T;Dunsby C;Neil MA;Roufosse CA;French PM
免疫荧光（IF）成像后的电子显微镜（EM）是诊断人类肾小球疾病的重要工具，但EM的实施仅限于专业机构，在许多国家都不可用。荧光显微镜的最新进展，使传统的宽视场荧光显微镜，以适应在适度的成本，以提供低于50纳米的生物标本的分辨率。我们表明，随机开关单分子定位显微镜可以应用于标准IF技术染色的临床组织学切片，这种超分辨IF可以提供一种替代手段来分辨超微结构，以帮助诊断EM不可用的肾脏疾病。我们已经实施了直接随机光学重建显微镜技术与临床批准的免疫荧光探针染色的基底层和免疫球蛋白G沉积的人肾活检冷冻切片。使用膜性肾小球肾炎，薄基底膜病变，狼疮性肾炎的情况下，我们比较这种方法的临床EM图像和增强成像相比，传统的IF显微镜。通过对临床冷冻肾活检的既定IF协议进行微小修改，我们相信传统宽视野显微镜的成本效益适应性可以广泛实施，以提供超分辨率图像信息，以帮助诊断人类肾小球疾病。
Electron microscopy (EM) following immunofluorescence (IF) imaging is a vital tool for the diagnosis of human glomerular diseases, but the implementation of EM is limited to specialised institutions and it is not available in many countries. Recent progress in fluorescence microscopy now enables conventional widefield fluorescence microscopes to be adapted at modest cost to provide resolution below 50 nm in biological specimens. We show that stochastically switched single‐molecule localisation microscopy can be applied to clinical histological sections stained with standard IF techniques and that such super‐resolved IF may provide an alternative means to resolve ultrastructure to aid the diagnosis of kidney disease where EM is not available. We have implemented the direct stochastic optical reconstruction microscopy technique with human kidney biopsy frozen sections stained with clinically approved immunofluorescent probes for the basal laminae and immunoglobulin G deposits. Using cases of membranous glomerulonephritis, thin basement membrane lesion, and lupus nephritis, we compare this approach to clinical EM images and demonstrate enhanced imaging compared to conventional IF microscopy. With minor modifications in established IF protocols of clinical frozen renal biopsies, we believe the cost‐effective adaptation of conventional widefield microscopes can be widely implemented to provide super‐resolved image information to aid diagnosis of human glomerular disease.
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DOI：
10.1073/pnas.0506010102
发表时间：
2005-12-06
影响因子：
11.1
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通讯作者：
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