A workflow for streamlined acquisition and correlation of serial regions of interest in array tomography.
A workflow for streamlined acquisition and correlation of serial regions of interest in array tomography.
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DOI：
10.1186/s12915-021-01072-7
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发表时间：
2021-07-30
期刊：
影响因子：
5.4
通讯作者：
Munck S
中科院分区：
文献类型：
--
作者：
Gabarre S;Vernaillen F;Baatsen P;Vints K;Cawthorne C;Boeynaems S;Michiels E;Vandael D;Gounko NV;Munck S
阵列层析成像是一种在细胞器水平上分辨精细细节的高分辨率成像方法，其优点是可以提供3D体积来显示组织环境。结合光学显微镜和电子显微镜(光镜、电子显微镜)技术，AT可以以一种相关的方式进行。然而，模式之间的相关性可能是一项挑战，在连续的切片中描绘特定的感兴趣区域可能会很耗时。集成光学和电子显微镜(ILEM)提供了提供良好相关性图像的可能性，并可能为相关AT提供理想的解决方案。在这里，我们报告一个在感兴趣区域之间自动导航的工作流。我们使用一种有针对性的方法，允许在不同尺度上成像特定的组织特征，如细胞器、细胞突起和细胞核，从而使用集成光学和电子显微镜(Ilem-AT)实现快速、直接相关的原位AT。我们的工作流程基于对初始透射光采集的截面边界的检测，该采集用作参考空间，以补偿截面之间的形状变化，并且随着放大倍数从LM增加到EM，我们应用逐步细化的局部化。通过最少的用户交互，这使得能够自主和快速地获取包含在不同放大倍数下关联的细胞和感兴趣的细胞器的区域，从而提供了一种更有效的获得3D图像的方式。我们提供了我们的方法和开发的软件工具的概念证明，使用高尔基体神经元浸染染色和细胞中荧光标记的蛋白质凝聚体。我们的方法便于在多个部分跟踪和重建蜂窝结构，目标是高分辨率ILEM，并且可以集成到现有设备中，无论是商业系统还是定制系统。网上版载有补充材料，可在10.1186/s12915-021-01072-7查阅。
Array tomography (AT) is a high-resolution imaging method to resolve fine details at the organelle level and has the advantage that it can provide 3D volumes to show the tissue context. AT can be carried out in a correlative way, combing light and electron microscopy (LM, EM) techniques. However, the correlation between modalities can be a challenge and delineating specific regions of interest in consecutive sections can be time-consuming. Integrated light and electron microscopes (iLEMs) offer the possibility to provide well-correlated images and may pose an ideal solution for correlative AT. Here, we report a workflow to automate navigation between regions of interest. We use a targeted approach that allows imaging specific tissue features, like organelles, cell processes, and nuclei at different scales to enable fast, directly correlated in situ AT using an integrated light and electron microscope (iLEM-AT). Our workflow is based on the detection of section boundaries on an initial transmitted light acquisition that serves as a reference space to compensate for changes in shape between sections, and we apply a stepwise refinement of localizations as the magnification increases from LM to EM. With minimal user interaction, this enables autonomous and speedy acquisition of regions containing cells and cellular organelles of interest correlated across different magnifications for LM and EM modalities, providing a more efficient way to obtain 3D images. We provide a proof of concept of our approach and the developed software tools using both Golgi neuronal impregnation staining and fluorescently labeled protein condensates in cells. Our method facilitates tracing and reconstructing cellular structures over multiple sections, is targeted at high resolution ILEMs, and can be integrated into existing devices, both commercial and custom-built systems. The online version contains supplementary material available at 10.1186/s12915-021-01072-7.
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DOI：
10.1023/b:visi.0000029664.99615.94
发表时间：
2004-11-01
影响因子：
19.5
作者：
Lowe, DG
通讯作者：
Lowe, DG
影响因子：
3
作者：
Faas, F. G. A.;Barcena, M.;Koster, A. J.
通讯作者：
Koster, A. J.
影响因子：
4.6
作者：
Chalfoun J;Majurski M;Blattner T;Bhadriraju K;Keyrouz W;Bajcsy P;Brady M
通讯作者：
Brady M
影响因子：
48
作者：
Conrad, Christian;Wuensche, Annelie;Tan, Tze Heng;Bulkescher, Jutta;Sieckmann, Frank;Verissimo, Fatima;Edelstein, Arthur;Walter, Thomas;Liebel, Urban;Pepperkok, Rainer;Ellenberg, Jan
通讯作者：
Ellenberg, Jan
影响因子：
4.6
作者：
Haring, Martijn T.;Liv, Nalan;Hoogenboom, Jacob P.
通讯作者：
Hoogenboom, Jacob P.