Versatile confocal microscopy for imaging objects oscillating at frequencies exceeding the frame rate

Abstract

Confocal microscopy typically has a low frame rate but allows high-resolution imaging deep within objects. We developed a versatile method to capture confocal images of periodically moving objects at frequencies exceeding the frame rate. This method requires no additional equipment or compromise in imaging capabilities and is compatible with all confocal laser scanning microscopes. We have demonstrated its use to visualize colloidal particle motion during elastic wave propagation at 1600 Hz—two orders of magnitude faster than standard confocal microscopy. Particle oscillation amplitude and phase exhibited attenuation and delay with propagation distance, allowing the determination of the elastic wave's amplitude attenuation and phase velocity. The single-particle analysis further revealed that the direction of elastic wave propagation depended on the orientation of the colloidal grains.