Extracellular filaments revealed by affinity capture cryo-electron tomography.

Area of Science:

• Cellular and Molecular Imaging
• Biophysics
• Microscopy Techniques

Background:

• Cryo-electron tomography (cryo-ET) offers nanoscale insights into cellular architecture.
• Micropatterning is vital for preparing adherent cells for cryo-ET workflows.
• Challenges exist in preparing minimally adherent cells for cryo-ET.

Purpose of the Study:

• To develop and demonstrate a micropatterning workflow for minimally adherent cell types using affinity capture.
• To enable high-resolution cryo-electron tomography (cryo-ET) and cryo-focused ion beam (cryo-FIB) imaging of T-cells and Jurkat cells.
• To improve workflow efficiency and cell positioning for cryo-ET data acquisition.

Main Methods:

• Development of an affinity capture system for cell micropatterning.
• Application of the workflow to human T-cells and Jurkat cells.
• Integration with cryo-focused ion beam (cryo-FIB) milling and cryo-electron tomography (cryo-ET) data acquisition.

Main Results:

• Successful nanoscale imaging of Jurkat cells, revealing extracellular filamentous structures.
• Demonstrated improved workflow efficiency with consistent production of well-positioned cells.
• Validated the utility of affinity capture for cryo-ET of minimally adherent cell types.

Conclusions:

• The developed affinity capture micropatterning workflow effectively prepares minimally adherent cells for cryo-ET.
• This method enhances imaging capabilities for studying cellular nanoscale architecture.
• The approach is adaptable for high-resolution imaging of diverse adherent and non-adherent cell types.