通过双视角斜平面显微镜进行高含量3D成像
Hugh Sparks1,2, Leo Rowe-Brown1,2, Yuriy Alexandrov1,2
1Light Community, Department of Physics, Imperial College London, South Kensington Campus, London SW7 2AZ, United Kingdom.
PNAS nexus
|December 8, 2025
概括
一个改进的双视角斜平面显微镜 (dOPM) 系统提供了高分辨率的3D成像与减少光漂白. 这种先进的光板光显微镜 (LSFM) 能够进行动态的活细胞和有机体研究.
科学领域:
- 生物物理学的生物物理.
- 显微镜的使用方法
- 光学工程是指光学工程.
背景情况:
- 斜平面显微镜 (OPM) 是一种光板光显微镜 (LSFM) 技术,使用单一的目标来激发和检测.
- 双视图OPM (dOPM) 是OPM的一个光学折叠变体.
研究的目的:
- 提供一个改进的dOPM系统,增强空间分辨率和光收集效率.
- 用不同的照明角度来评估系统性能,并将实验数据与模拟进行比较.
- 为了展示系统的实时成像,多视场3D成像和生物样本中的动态事件观测的能力.
主要方法:
- 开发了一种改进的dOPM系统,使用60×/1.2NA水浸镜头.
- 在35°和45°的照明角度测量空间分辨率 (FWHM) 和光学切割强度.
- 将光收集效率与矢量射线追踪模拟进行比较.
- 在凝中进行的有机体和细胞的时间延迟成像,包括FUCCI细胞周期报告员量化.
- 在96个井板中对生物样本进行了多视场成像.
主要成果:
- 35°的照明角度带来了稍微更好的侧面分辨率和收集效率.
- 合并的dOPM视图实现了0.29μm (x),0.31μm (y) 和0.83μm (z) 的中珠FWHM,光学截面为2.45-3.00μm.
- 与活体有机体的广场光成像相比,dOPM的光漂白减少.
- 在原凝中成功成像细胞,量化了FUCCI报告员在球体中药物剂量反应曲线.
- 观察到的动态生物事件,包括有机体光动态和ex vivo组织切片中的迁移.
结论:
- 改进的dOPM系统提供了高分辨率,低光损伤的3D成像功能.
- dOPM适用于动态生物过程的时间间隔研究和多视场查.
- 该系统可在各种生物环境中对细胞和有机体行为进行定量分析.
相关概念视频
Three-Dimensional Microscopy in Microbiology
741
Three-dimensional imaging techniques are essential in cell biology, allowing researchers to visualize intricate cellular structures with high resolution. Two prominent methods, Differential Interference Contrast Microscopy (DIC) and Confocal Scanning Laser Microscopy (CSLM), provide distinct advantages for imaging live and thick specimens, respectively.Differential Interference Contrast MicroscopyDIC microscopy enhances contrast in transparent, unstained samples by converting phase...
741
Two-Dimensional Microscopy in Microbiology
1.0K
Two-dimensional (2D) microscopy encompasses a range of optical techniques that capture images within a single focal plane, offering detailed representations of microscopic structures. These techniques are essential in biological and medical research, enabling the visualization of cellular and subcellular structures with different levels of contrast and specificity.There are several major types of 2D microscopy, each with strengths and applications.Bright-Field MicroscopyBright-field microscopy...
1.0K
Confocal Fluorescence Microscopy
19.9K
Confocal microscopy is an advanced microscopic technique. The prime advantage of the confocal microscope over other microscopy techniques is its ability to block the out-of-focus light from the illuminated samples using pinholes. It is widely used with fluorescence optics to obtain high-resolution, sharp contrast images. Unlike optical microscopes, confocal microscopes use a focused beam of light laser to scan the entire sample surface at different z-planes. These microscopes are, therefore,...
19.9K
Electron Microscope Tomography and Single-particle Reconstruction
2.8K
Transmission electron microscopy (TEM) can be used to determine the 3D structure of biological samples with the help of techniques such as electron microscope tomography and single-particle reconstruction. While single-particle reconstruction can examine macromolecules and macromolecular complexes in vitro conditions only, tomography permits the study of cell components or small cells in vivo.
Electron Tomography
Electron tomography can be performed either in TEM or STEM (scanning transmission...
Electron Tomography
Electron tomography can be performed either in TEM or STEM (scanning transmission...
2.8K
Imaging Biological Samples with Optical Microscopy
8.7K
Optical microscopy uses optic principles to provide detailed images of samples. Antonie van Leeuwenhoek designed the first compound optical microscope in the 17th century to visualize blood cells, bacteria, and yeast cells. In 1830, Joseph Jackson Lister created an essentially modern light microscope. The 20th century saw the development of microscopes with enhanced magnification and resolution.
In optical microscopy, the specimen to be viewed is placed on a glass slide and clipped on the stage...
In optical microscopy, the specimen to be viewed is placed on a glass slide and clipped on the stage...
8.7K
Phase Contrast and Differential Interference Contrast Microscopy
11.9K
Phase-Contrast Microscopes
In-phase-contrast microscopes, interference between light directly passing through a cell and light refracted by cellular components is used to create high-contrast, high-resolution images without staining. It is the oldest and simplest type of microscope that creates an image by altering the wavelengths of light rays passing through the specimen. Altered wavelength paths are created using an annular stop in the condenser. The annular stop produces a hollow cone of...
In-phase-contrast microscopes, interference between light directly passing through a cell and light refracted by cellular components is used to create high-contrast, high-resolution images without staining. It is the oldest and simplest type of microscope that creates an image by altering the wavelengths of light rays passing through the specimen. Altered wavelength paths are created using an annular stop in the condenser. The annular stop produces a hollow cone of...
11.9K


