Mojiri, S. | bioRxiv Science

bioRxiv2021

Instant three color multi-plane fluorescence microscopy

One of the most widely used microscopy techniques in biology and medicine is fluorescence microscopy, offering high specificity in labeling as well as maximum sensitivity. For live cell imaging, the ideal fluorescence microscope should offer high spatial resolution, fast image acquisition, three-dimensional sectioning, and multi-color detection. However, most existing fluorescence microscopes have to compromise between these different requirements. Here, we present a multi-plane multi-color wide-field microscope that uses a dedicated beam-splitter for recording volumetric data in eight focal planes and for three emission colors with frame rates of hundreds of volumes per second. We demonstrate the efficiency and performance of our system by three-dimensional imaging of multiply labeled fixed and living cells.

biophysics↗

bioRxiv2020

Three-dimensional beating dynamics of Chlamydomonas flagella

Axonemes are the basic structure of motile cilia and flagella, and the investigation of how they function and move requires rapid three-dimensional imaging. We built a multi-plane phase-contrast microscope for imaging the three-dimensional motion of unlabeled flagella of the model organism Chlamydomonas reinhardtii with sub-m spatial and 4 ms temporal resolution. This allows us to observe not only bending but also the three-dimensional torsional dynamics of these small structures. We observe that flagella swim counter-clockwise close to a surface, with negatively-valued torsion at their basal and positively-valued torsion at their distal tips. To explain the torsional dynamics and signature, we suggest the existence of an intrinsic negative twist at the basal end that is untwisted by active positive-twist-inducing dynein motor proteins. Moreover, dyneins walking towards the basal induce an opposite twist at the distal tip. Bending of the whole axoneme structure then translates this twist into an observable torsion. This interconnection between chiral structure, twist, curvature, and torsion is fundamental for understanding flagellar mechanics.

biophysics↗