For decoding the mechanism of how cells and organs function information on their ultrastructure is essential. High-resolution 3D imaging has revolutionized morphology. Serial block face scanning electron microscopy (SBF-SEM) offers non-laborious, automated imaging in 3D of up to ~ 1 mm3 large biological objects at nanometer-scale resolution. For many samples there are obstacles. Quality imaging is often hampered by charging effects, which originate in the nonconductive resin used for embedding. Especially, if the imaged region of interest (ROI) includes the surface of the sample and neighbours the empty resin, which insulates the object. This extra resin also obscures the sample’s morphology, thus making navigation to the ROI difficult. Using the example of small arthropods and a fish roe we describe a workflow to prepare samples for SBF-SEM using the minimal resin (MR) embedding method. We show that for imaging of surface structures this simple approach conveniently tackles and solves both of the two major problems—charging and ROI localization—that complicate imaging of SBF-SEM samples embedded in an excess of overlying resin. As the surface ROI is not masked by the resin, samples can be precisely trimmed before they are placed into the imaging chamber. The initial approaching step is fast and easy. No extra trimming inside the microscope is necessary. Importantly, charging is absent or greatly reduced meaning that imaging can be accomplished under good vacuum conditions, typically at the optimal high vacuum. This leads to better resolution, better signal to noise ratio, and faster image acquisition. In MR embedded samples charging is minimized and ROI easily targeted. MR embedding does not require any special equipment or skills. It saves effort, microscope time and eventually leads to high quality data. Studies on surface-linked ROIs, or any samples normally surrounded by the excess of resin, would benefit from adopting the technique.

中文翻译：

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SBF-SEM 样品的最小树脂嵌入减少了充电并有助于寻找表面连接的感兴趣区域

为了解码细胞和器官如何发挥作用的机制，有关其超微结构的信息至关重要。高分辨率 3D 成像彻底改变了形态学。串行块面扫描电子显微镜 (SBF-SEM) 可以以纳米级分辨率对高达约 1 mm3 的大型生物物体进行轻松、自动的 3D 成像。对于许多样品来说都存在障碍。高质量成像常常受到充电效应的影响，这种效应源于用于嵌入的非导电树脂。特别是，如果成像的感兴趣区域 (ROI) 包括样品表面并与绝缘物体的空树脂相邻。这种额外的树脂还会模糊样品的形态，从而使导航到 ROI 变得困难。以小型节肢动物和鱼子为例，我们描述了使用最小树脂 (MR) 嵌入方法制备 SBF-SEM 样品的工作流程。我们表明，对于表面结构成像，这种简单的方法可以方便地处理和解决两个主要问题——充电和 ROI 定位——这使得嵌入过量树脂中的 SBF-SEM 样品的成像变得复杂。由于表面 ROI 未被树脂遮盖，因此可以在将样品放入成像室之前对其进行精确修整。最初的接近步骤快速而简单。显微镜内部不需要额外的修剪。重要的是，不存在充电或大大减少充电意味着可以在良好的真空条件下（通常在最佳高真空下）完成成像。这会带来更好的分辨率、更好的信噪比和更快的图像采集。在 MR 嵌入式样品中，收费被最小化，并且可以轻松实现投资回报率目标。MR 嵌入不需要任何特殊设备或技能。它节省了精力和显微镜时间，并最终获得高质量的数据。对表面连接的 ROI 或任何通常被过量树脂包围的样品的研究将受益于采用该技术。