Development of Planar Illumination Strategies for Solving Mysteries in the Sub-Cellular Realm

Abstract

Optical microscopy has vastly expanded the frontiers of structural and functional biology, due to the non-invasive probing of dynamic volumes in vivo. However, traditional widefield microscopy illuminating the entire field of view (FOV) is adversely affected by out-of-focus light scatter. Consequently, standard upright or inverted microscopes are inept in sampling diffraction-limited volumes smaller than the optical system’s point spread function (PSF). Over the last few decades, several planar and structured (sinusoidal) illumination modalities have offered unprecedented access to sub-cellular organelles and 4D (3D + time) image acquisition. Furthermore, these optical sectioning systems remain unaffected by the size of biological samples, providing high signal-to-noise (SNR) ratios for objective lenses (OLs) with long working distances (WDs). This review aims to guide biologists regarding planar illumination strategies, capable of harnessing sub-micron spatial resolution with a millimeter depth of penetration.

A young researcher’s guide to three-dimensional fluorescence microscopy of living cells

“…While light-sheet microscopy has been used for more than 20 years to image larger samples under low magnification, more recently it has been adapted to imaging single cells with diffraction-limited spatial resolution (19). The most successful variant of high-magnification lightsheet microscopy is the lattice light-sheet microscopy, which uses sophisticated optoelectronic techniques to generate a thin and stable sheet of light to illuminate and image only a single section of the specimen at a time (20,21).…”

Scale space detector for analyzing spatiotemporal ventricular contractility and nuclear morphogenesis in zebrafish

“…Conventionally, invasive sectioning procedures have offered revolutionary insights into aberrant tissue up to the cellular scale ( Bensley et al., 2016 ; Javaeed et al., 2021 ; Teranikar et al., 2022 ). However, histopathological analysis currently suffers from severe limitations, primarily disruption to tissue homeostasis ( Kleczek et al., 2020 ; Teranikar et al., 2022 ).…”

“…Conventionally, invasive sectioning procedures have offered revolutionary insights into aberrant tissue up to the cellular scale ( Bensley et al., 2016 ; Javaeed et al., 2021 ; Teranikar et al., 2022 ). However, histopathological analysis currently suffers from severe limitations, primarily disruption to tissue homeostasis ( Kleczek et al., 2020 ; Teranikar et al., 2022 ). With respect to these drawbacks, the optical sectioning modality light sheet fluorescence microscopy (LSFM) has proved instrumental in probing dynamic organogenesis several millimeters inside tissue.…”