BPS2026 – An automated mechanical stretcher with integrated fiducial tracking software for monitoring real-time cell expansion

Conventional expansion microscopy (ExM) can increase the effective resolution of imaging using water uptake to expand a sample above the diffraction limit; however, osmotic ExM involves labor-intensive protocols, requires fixation that can lead to physical distortions to sample, and signal degradation. To overcome these challenges, we have developed a novel electromechanical platform that physically expands cells using tensile force to achieve high-resolution imaging. Our tensile ExM system integrates an iris-based mechanical stretcher controlled using a stepper motor and micro-controller chip along with a Python-automated data acquisition to apply precise, uniform tensile force to cells cultured on a stretchable hydrogel. Here, we detail our system design, electronics, and high-fidelity feature tracking of nano-sized fiducial markers within the hydrogel during expansion using optical microscopy. The iris-expansion device was fabricated using cost-effective 3D printed and laser cut components. PCB-controlled stepper motor drives a belt to actuate the tensile ExM device arms, expanding the hydrogel sample held by grippers. A Python script automates the experiment by controlling the expansion system via a microcontroller and executing an image processing workflow. The workflow uses a gradient-based software autofocus with a Tenengrad function, image segmentation, and a nearest-neighbor algorithm to track fiducial markers, which eliminates manual focusing and thereby improves efficiency. Overall, our iris tensile expansion system provides a robust framework for investigating the real-time dynamics of cellular components under mechanical stress, yielding new insights into fundamental biophysical questions.