Laboratory-based cryogenic soft x-ray tomography with correlative cryo-light and electron microscopy

Area of Science:

• Cell Biology
• Microscopy
• Biophysics

Background:

• Understanding cellular ultrastructure is crucial for biological research.
• Traditional methods like electron microscopy require chemical fixation and sectioning, which can introduce artifacts.
• Cryo-soft X-ray microscopy offers a potential alternative for imaging hydrated, unstained cells.

Purpose of the Study:

• To present a novel laboratory-based cryogenic soft X-ray microscope.
• To demonstrate its capability for whole cell tomography of frozen hydrated samples.
• To enable correlative imaging with other cryo-microscopy techniques.

Main Methods:

• Development of a compact cryogenic soft X-ray microscope.
• Whole cell tomography of Saccharomyces cerevisiae.
• Imaging unstained yeast cells using cryo-fluorescence light microscopy, cryo-soft X-ray microscopy, and cryo-transmission electron microscopy.

Main Results:

• Achieved better than 50 nm half-pitch spatial resolution.
• Visualized internal subcellular structures of yeast cells.
• Detected structures as small as 90 nm with a low radiation dose (7.2 MGy).
• Enabled imaging of biological cells up to 5 μm thick.

Conclusions:

• The developed cryogenic soft X-ray microscope provides high-resolution 3D ultrastructure determination of whole cells.
• It allows multimodal correlative imaging of the same cell.
• This technology offers greater access to imaging unstained, hydrated cells without fixation or sectioning.