Biology subjects

Oketani, R.

Publications and source records attributed to Oketani, R..

4 recordsLinked to original sources

bioRxiv2026

Long-term single-particle tracking by NIR imaging using Au42 (gold) quantum needles

Single-particle tracking (SPT) over time enables direct observation of molecular transport and interactions in living cells. Fluorescence-based SPT has provided insights into intracellular processes such as endocytosis, receptor signaling, and drug delivery. Extending the observation window to several hours and beyond is critical for capturing slow intracellular dynamics, including the full course of endosomal trafficking, the long-term accumulation of particles within subcellular compartments, and transitions between transport modes that occur on hour-scale timescales. However, long-term intracellular SPT under visible-wavelength excitation remains challenging because fluorescence probes generally suffer from photobleaching and phototoxicity. While near-infrared (NIR) excitation can simultaneously mitigate these issues, generally weak emission of NIR-emitting dyes has hindered its wide application in long-term SPT. Here, we demonstrate long-term NIR SPT using atomically precise gold quantum needles, Au42(PET)32 (PET = 2-phenylethanethiolate). Continuous tracking of intracellular particles in living HEK293 cells was achieved for up to 12 h. Trajectory analysis revealed temporal transitions between directional and diffusive transport, as well as the accumulation of multiple particles within localized intracellular domains over several-hour timescales. The high photostability of Au42, combined with low phototoxicity of NIR excitation, enables visualization of intracellular transport dynamics over timescales difficult to access using conventional visible fluorescent probes. These results establish Au42-based NIR imaging as a platform for long-term, low-phototoxicity intracellular SPT and provide a framework for investigating slow intracellular dynamics in living systems.

biophysics↗

bioRxiv2025

Broadband electronic resonance coherent anti-Stokes/Stokes Raman scattering microscopy

Coherent Raman imaging (CRI) enables label-free chemical imaging based on intrinsic molecular vibrations, but its applicability is often limited by low sensitivity, hindering the detection of low-abundance biomolecules. While electronic resonance significantly enhances sensitivity, most resonance CRI implementations rely on narrowband excitation and/or detection, which limits spectral coverage and makes it difficult to distinguish target molecules from backgrounds. Here we address these challenges by developing broadband electronic-resonance coherent anti-Stokes/Stokes Raman scattering (BER-CARS/CSRS) microscopy. We show that BER-CARS/CSRS enables highly sensitive, label-free imaging of endogenous chromophores with broad spectral coverage of the entire fingerprint region. Specifically, we captured time-lapse images across the fingerprint region, visualizing low-abundance cytochromes alongside abundant biomolecules (lipids, proteins, nucleic acids) in living HEK293 cells. Furthermore, we applied the method to complex biological tissues, mapping distinct distributions of cytochromes in mouse brain slices, highlighting their characteristic localizations in the cortex and the cerebral ventricle wall. Our results demonstrate that BER-CARS/CSRS is a powerful platform for highly sensitive chemical imaging, from large-area tissue mapping to organelle-level dynamics. By coupling resonance enhancement with broadband fingerprinting, BER-CARS/CSRS enables dynamic, label-free phenotyping of mitochondrial and physiological states from single cells to tissues, opening a path to quantitative, slide-scale chemical histopathology and intraoperative margin assessment.

bioengineering↗

bioRxiv2024

Label-free visualization of ciliary rootlets in mouse brain

Neuronal primary cilia are important role in brain development, sensory perception and neurogenesis. Rootletin, a fibrous protein composed of coiled-coil motifs, is a major structural component of ciliary rootlets and is essential for understanding ciliary functions. However, the precise mechanisms by which Rootletin influences ciliary dynamics and impacts neuronal function remain largely unknown, primarily due to the challenges in visualizing these structures. Here, we describe a label-free, rapid, and highly sensitive method to visualize Rootletin molecules in brain tissue. This platform integrates a second harmonic generation (SHG) microscope and background reduction by a one-step chemical pretreatment. Additionally, we employ coherent anti-Stokes Raman scattering imaging to simultaneously determine the cellular regions and intracellular locations of SHG signals. By applying this multimodal multiphoton imaging to mouse hippocampus, we found that neuronal ciliary rootlets were found to exhibit highly organized specific intracellular distributions. Moreover, the formation of ciliary rootlets precedes that of primary cilia. These findings highlight the utility of our label-free imaging platform in developmental and neuroscience research, providing a new tool to characterize ciliary dynamics and neuronal function.

biophysics↗

bioRxiv2023

Selective plane activation structured illumination microscopy

Resolution enhancement in structured illumination microscopy is hindered when volumetric samples are observed owing to background signals from out-of-focus planes. In this study, we utilized selective plane illumination and reversibly photoswitchable fluorescent proteins to realize structured illumination within the focal plane and eliminate the out-of-focus background. We demonstrated high-resolution fluorescence imaging of volumetric samples, including dense fluorescent objects in a cell spheroid.

biophysics↗