Tensile expansion microscopy applies mechanical force to super-resolve fixed and image live cellular samples

Area of Science:

• Biophysics
• Cell Biology
• Microscopy

Background:

• Biophysical studies require techniques with biologically relevant spatial and temporal scales.
• Traditional Expansion Microscopy (ExM) offers super-resolution but limits imaging to pre- and post-expansion states, hindering dynamic observations.
• ExM limitations include fragmentation, deformation, signal loss, and the need for chemical fixation, impacting reproducibility and dynamic studies.

Purpose of the Study:

• To develop a novel microscopy technique enabling mechanical expansion of both fixed and live cellular samples.
• To overcome limitations of osmotic ExM, such as restricted imaging windows and fixation requirements.
• To enable real-time monitoring of cellular dynamics at enhanced spatial resolutions.

Main Methods:

• Developed Tensile Expansion Microscopy (TExM) using stretchable alginate-Ca2+/polyacrylamide hydrogels and an electromechanical iris expansion device.
• Incorporated two-photon polymerized microscale fluorescent fiducial markers for tracking sample distortion during expansion.
• Applied TExM to fixed NIH 3T3 fibroblasts and live HeLa cells with internal fluorescent reporters.

Main Results:

• Achieved controllable and repeatable hydrogel expansion up to 3.3× with minimal distortion (<12 µm across 1.3 mm²).
• Obtained super-resolution imaging of fixed cells with 100 nm resolution for microtubules.
• Demonstrated live cell imaging showing increased cell size and separation under 3.2× linear expansion.

Conclusions:

• TExM enables continuous, stepwise, and precise temporal modulation of substrate strain for real-time dynamics monitoring.
• This technique provides higher spatial resolution for both fixed and live cellular processes.
• TExM's compatibility with other imaging methods broadens its applicability for biophysical research.