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

Nadav Scher (1) , Katya Rechav (1) , Perrine Paul-Gilloteaux (2) , Ori Avinoam (1)

1 Weizmann Institute of Science [Rehovot, Israël]
2 Nantes Univ - SFR François Bonamy - Structure fédérative de recherche François Bonamy

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).

Mots clés: Cell Biology; Cryo-EM; Microscopy

Domaines: Traitement du signal et de l'image [eess.SP]; Biologie cellulaire; Imagerie

Nadav Scher, Katya Rechav, Perrine Paul-Gilloteaux, Ori Avinoam. Sample preparation and image registration for correlative cryo-FM and cryo-FIB-SEM of plunge-frozen mammalian cells. STAR Protocols, 2022, 3 (1), pp.101142. ⟨10.1016/j.xpro.2022.101142⟩. ⟨hal-03745072⟩