A cryo-CLEM–guided workflow for quantitative assessment and optimization of cryo-ET specimen quality
Year: 2026
Type: article
Abstract: Cryo-correlative light and electron microscopy (cryo-CLEM) is an essential technique for precisely targeting regions of interest in vitrified biological specimens for in situ cellular cryo-electron tomography (cryo-ET). Despite its widespread use, the potential of cryo-CLEM remains underexploited because its imaging performance under cryogenic conditions has not been characterized. We evaluated the imaging capabilities of 2 commercially available cryo-CLEM systems-a stand-alone Leica EM cryo-CLEM microscope and the integrated Thermo Fisher Scientific iFLM coupled to a cryo-focused ion beam platform-by quantifying their spatial resolution and signal-to-noise ratio using vitrified cellular specimens expressing green fluorescent protein (GFP)-labeled proteins. The EM cryo-CLEM setup provided a higher signal-to-noise ratio and faster acquisition via tile imaging, whereas the iFLM offered a wider field of view, revealing complementary strengths between the 2 instruments. Furthermore, we established a quantitative correlation between fluorescence intensity and lamella thickness, demonstrating that cryo-CLEM imaging can serve as a practical indicator of lamella quality during early specimen preparation, before cryo-ET data collection. Collectively, our results show that cryo-CLEM not only facilitates precise targeting but also provides quantitative feedback for optimizing lamella quality, thereby streamlining the overall cryo-ET workflow.
Source: Molecules and Cells
Authors: Minjung Kim, Junsun Park, Soung-Hun Roh
Institution: Seoul National University
Language: English
Subfield: Structural Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $2,500
OpenAlex ID: W7129058741