Advances in Cryo-focused Ion Beam-Scanning Electron Microscopy Imaging Technology

doi:10.11983/CBB21161

Abstract

Abstract: Cryo-focused ion beam-scanning electron microscopy (Cryo-FIB-SEM) is an emerging technology designed for advanced imaging detection, which performs in situ by combining Cryo-FIB milling and Cryo-SEM imaging, and has facilitated the visualization of the native structures of biological sample in the context of the cellular environment in the frozen hydrated state. In recent years, a series of important advances have been achieved in the application of this technology in the research field of life science. In this review, we summarize its application in cryo-volume serial imaging, and in combination with cryo-correlative light and electron microscopy (CLEM), cryo-transmission SEM (TSEM), cryo-lamella preparation monitoring, and Cryo-SEM image processing. We also provide future prospective on future development and application of Cryo-FIB-SEM in three-dimensional in situ imaging of large volume biological samples.

Key words: cryo-focused ion beam-scanning electron microscopy (Cryo-FIB-SEM), cryo-correlative light and electron microscopy (Cryo-CLEM), cryo-volume serial imaging, cryo-transmission scanning electron microscopy imaging

Xing Jia, Fei Sun, Gang Ji. Advances in Cryo-focused Ion Beam-Scanning Electron Microscopy Imaging Technology[J]. Chinese Bulletin of Botany, 2022, 57(1): 24-29.

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URL: https://www.chinbullbotany.com/EN/10.11983/CBB21161

https://www.chinbullbotany.com/EN/Y2022/V57/I1/24

Figures/Tables 3

Figure 1 Cryo-FIB-SEM application fields Cryo-FIB-SEM application fields contain cryo-volume serial imaging, cryo-correlative light and electron microscopy imaging, cryo-transmission scanning electron microscopy imaging, cryo-lamella preparation monitoring, and cryo-SEM image processing.

Table 1 Summary of serial Cryo-FIB-SEM parameters

	FIB束流(pA)	SEM束流(pA)	SEM电压(kV)	剂量 pA×nS/Å²	驻留时间(ns)	平均线数	像素尺寸x-y-z (nm)	图像尺寸x-y (像素)	切片数	总体积 (μm³)	耗时 (h)
对照	700	59	1.9	22.4	200	19	10×10×20	3072×1150	575	4062	5.5
患者	700	35	2.3	21.3	100	61	10.5×10.5×21	4096×3072	2018	58789	17.5

Figure 2 Experimental workflow including multiple correlative imaging steps under cryogenic conditions via different imaging platforms

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