High-fidelity microsecond-scale cellular imaging using two-axis compressed streak imaging fluorescence microscopy.

Compressed streak imaging (CSI) is a computational imaging strategy that can acquire video at over 150 trillion frames per second. Despite this achievement, CSI faces challenges in detecting subtle intensity fluctuations in slow-moving, continuously illuminated objects. This limitation, largely attributable to high streak compression and motion blur, has curtailed the broader adoption of CSI in cellular fluorescence microscopy. To address these issues and expand the utility of CSI, we developed a two-axis compressed streak imaging (TACSI) method that results in significant improvements to the reconstructed video fidelity. TACSI introduces a second scanning axis which shuttles a conjugate image of the object with respect to the coded aperture. The moving image decreases the streak compression ratio and produces a "flash and shutter" phenomenon that reduces coded aperture motion blur, overcoming the limitations of current CSI technologies. This approach is supported with an analytical model describing the TACSI compression ratio, along with simulated and empirical measurements. We demonstrate TACSI's ability to measure rapid variations in cell membrane potentials, previously unattainable with conventional CSI. This method has broad implications for high-speed photography, including visualization of action potentials, muscle contractions, and enzymatic reactions that occur on microsecond and faster timescales using fluorescence microscopy.