%0 Journal Article
%T A protocol for cryogenic volumetric imaging using serial plasma FIB/SEM.
%A Dumoux M
%A Smith JLR
%A Glen T
%A Grange M
%A Darrow MC
%A Naismith JH
%J Methods Cell Biol
%V 177
%N 0
%D 2023
%M 37451772
%F 1.829
%R 10.1016/bs.mcb.2023.01.015
%X 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.