Sample preparation and image registration for correlative cryo-FM and cryo-FIB-SEM of plunge-frozen mammalian cells

Abstract

Summary: We recently demonstrated how lipid droplets can serve as in situ fiducials for correlating cryo-fluorescence microscopy (cryo-FM) and cryo-focused ion beam scanning electron microscopy (cryo-FIB-SEM) datasets of mammalian cells grown on grids. Here we describe a step-by-step protocol for correlative cryo-FM and cryo-FIB-SEM, starting from sample preparation of C2C12 cell line, followed by imaging with cryo-FM and cryo-FIB-SEM. Finally, we detail how to perform the 3D-correlation with sub-micron accuracy.For complete details on the use and execution of this profile, please refer to Scher et al. (2021).

Keywords

• Cell Biology
• Microscopy
• Cryo-EM