Multi-resolution correlative focused ion beam scanning electron microscopy: Applications to cell biology

作者：Narayan Kedar; Danielson Cindy M; Lagarec Ken; Lowekamp Bradley C; Coffman Phil; Laquerre Alexandre; Phaneuf Michael W; Hope Thomas J; Subramaniam Sriram^*

来源：Journal of Structural Biology, 2014, 185(3): 278-284.

DOI：10.1016/j.jsb.2013.11.008

• Three-dimensional electron microscopy
• Ion abrasion scanning electron microscopy
• Correlative microscopy
• HIV-1 core
• 3D imaging of bacteria
• Tomography
• FIB-SEM

Efficient correlative imaging of small targets within large fields is a central problem in cell biology. Here, we demonstrate a series of technical advances in focused ion beam scanning electron microscopy (FIB-SEM) to address this issue. We report increases in the speed, robustness and automation of the process, and achieve consistent z slice thickness of similar to 3 nm. We introduce "keyframe imaging" as a new approach to simultaneously image large fields of view and obtain high-resolution 3D images of targeted sub-volumes. We demonstrate application of these advances to image post-fusion cytoplasmic intermediates of the HIV core. Using fluorescently labeled cell membranes, proteins and HIV cores, we first produce a "target map" of an HIV infected cell by fluorescence microscopy. We then generate a correlated 3D EM volume of the entire cell as well as high-resolution 3D images of individual HIV cores, achieving correlative imaging across a volume scale of 10(9) in a single automated experimental run.

• 出版日期2014-3
• 单位西北大学