Rapid multi-plane phase-contrast microscopy reveals torsional dynamics in flagellar motion

High speed volumetric optical microscopy is an important tool for observing rapid processes in living cells or for real-time tracking of sub-cellular components. However, the 3D imaging capability often comes at the price of a high technical complexity of the imaging system and/or the requirement of demanding image analysis. Here, we propose a combination of conventional phase-contrast imaging with a customized multi-plane beam-splitter for enabling simultaneous acquisition of images in eight different focal planes. Our method is technically straightforward and does not require complex post-processing image analysis. We apply our multiplane phase-contrast microscope to the real-time observation of the fast motion of reactivated Chlamydomonas axonemes with sub-µm spatial and 4 ms temporal resolution. Our system allows us to observe not only bending but also the three-dimensional torsional dynamics of these micro-swimmers.