Cell Biology

Many interesting cellular structures are smaller than the optical diffraction limit of ~200-300 nm. These include the substructures of most organelles and all macromolecular machines, channels, and receptors.

Asking and answering questions about the molecular organization, interactions, and stoichiometry of these and other structures requires imaging of labeled structures at resolutions below the diffraction limit of light.

Conventional Microscopy Techniques in Cell Biology

Conventional light microscopy methods, like widefield and confocal microscopy, can image and identify specifically labeled cellular structures. However, they are limited in their ability to image below the optical diffraction limit. Alternatively, electron microscopy (EM) can achieve resolutions down to 0.1-0.2 nm laterally, but specific labeling or quantitative measurement of molecules within the cell is very challenging.

细胞生物学的超分辨率荧光显微镜

超分辨率显微镜支持高分辨率成像 - 横向下至20 nm，沿光轴50 nm - 专门标记的细胞结构。因此，它弥合了常规光与电子显微镜（EM）技术之间的差距。

_{*右：用Alexa 647对TOM20染色的线粒体，用SMLM（DSTORM）成像。}

The high resolution (20 nm laterally) paired with the ability to image specifically labeled structures makes SMLM a powerhouse solution in cell biology research. Discoveries made with SMLM include the visualization of local movements of chromatin domains in HeLa cells, the eightfold symmetry within the nuclear pore complex of Xenopus oocytes and the radial ninefold symmetry of the centriolar protein CEP164, and the organization of connected tubular structures within the endoplasmic reticulum[1, 2].

Examples of Nanoscale Cellular Structures Suitable for SMLM Research

超分辨率显微镜支持高分辨率成像 - 横向下至20 nm，沿光轴50 nm - 专门标记的细胞结构。因此，它弥合了常规光与电子显微镜（EM）技术之间的差距。

在确定SMLM是否是正确的解决方案时，必须考虑可视化感兴趣结构所需的分辨率。提供了一些适合用SMLM成像的衍射限制结构大小的示例。示例*包括（但不限于）：

• 核孔复合物的扩散通道（长度为40-50nm）
• Nucleoid in mitochondria (~100 nm in diameter)
• Microtubules (24 nm in thickness)
• 核糖体（直径约20-30 nm）
• Bacterial heat shock protein DegP (20 nm in diameter)
• 圆柱形淀粉核涂布les in the chloroplast (107 ± 26 nm in diameter)
• 小鼠细胞中的内吞坑（直径86±2.4nm）
• 大鼠肝线粒体（直径30-40 nm）中Cristae的管状片段
• 质膜小窝（直径50-100 nm）
• 细菌鞭毛的钩子（长度约55 nm）

_{*Examples of nanoscale substructures and their dimensions are from 'The database of Useful Biological Numbers'. Link:https://bionumbers.hms.harvard.edu/search.aspx}

在超分辨率显微镜方法中，单分子定位显微镜（SMLM）可以说是最适合细胞生物学研究的。

具体来说，SMLM is the best method for studying specific nanoscale structures within cells when the research question requires:

• 可视化分子相距至20 nm；
• Labeling of multiple specific molecular structures; or
• 定量数据收集在此规模上。
  

SMLM is already being used to answer previously unexplored questions in cell biology. SMLM is ideal for imaging multiple specific structures in 3D and at high resolution, as well as quantifying data. See the sample data and descriptions, below, to learn more about research applications that can be uniquely achieved with SMLM:

固定样品成像

Microtubules acquired on the VXL - dSTORM images of fixed microtubules labeled with Alexa 647 using primary/seconday antibody labeling.

Live Cell Imaging

Acquired on the VXL - Imaging of live BSC1 cells labeled with Alexa 647 transferrin.

布鲁克's Vutara VXL offers best-in-class ease of use and imaging depth. Key features supporting the needs of cell biology researchers include:

• 自动化工作流：SRX软件使得可以轻松获取必要的数据集，并且工作流程使用户可以专注于生物学问题，而不是修补复杂的显微镜设置。
• Superior 3D Imaging and Sample Flexibility:专有双平面检测提供了三维信息，并在组织切片中进行成像。
• 无限的多路复用：The integrated Microfluidics Unit allows sequential labeling of an unlimited number of fluorescent probes.
• Expert Applications Support:Vutara VXL users can receive personalized guidance for sample prep optimization specific to their research objectives.