Abstract：
Cryogenic volumetric imaging using serial plasma focused ion beam scanning electron microscopy (serial pFIB/SEM) is a new and exciting correlative volume electron microscopy (vEM) technique. It enables visualization of un-stained, cryogenically immobilized cells and tissues with ∼20-50nm resolution and a field of view of ∼10-30μm resulting in near-native state imaging and the possibility of microscale, mesoscale and nanoscale correlative imaging. We have written a detailed protocol for optimization of FIB and SEM parameters to reduce imaging artefacts and enable downstream computational processing and analysis. While our experience is based on use of a single system, the protocol has been written to be as hardware and software agnostic as possible, with a focus on the purpose of each step rather than a fully procedural description to provide a useful resource regardless of the system/software in use.
摘要：
使用串行等离子体聚焦离子束扫描电子显微镜（串行pFIB/SEM）进行低温体积成像是一种新颖且令人兴奋的相关体积电子显微镜（vEM）技术。它可以实现未染色的可视化，低温固定的细胞和组织，分辨率为20-50nm，视野为10-30μm，导致近天然状态成像和微尺度的可能性，中尺度和纳米级相关成像。我们已经为FIB和SEM参数的优化编写了详细的协议，以减少成像伪影并实现下游计算处理和分析。虽然我们的经验是基于单一系统的使用，协议已被编写为尽可能与硬件和软件无关，专注于每个步骤的目的，而不是完整的过程描述，以提供有用的资源，而不管使用的系统/软件如何。
