Journal Article

Quantitative 3D imaging of whole, unstained cells by using X-ray diffraction microscopy

Ishikawa, Tetsuya, Chen, Chien-Chun, Lu, Chien-Hung, Miao, Jianwei, Tamanoi, Fuyuhiko, Urano, Jun, Lee, Ting-Kuo, Nakashima, Akio, Murnane, Margaret M., Song, Changyong, Jiang, Huaidong, Xu, Rui, Raines, Kevin S., Fahimian, Benjamin P.

2010

Overview

Microscopy has greatly advanced our understanding of biology. Although significant progress has recently been made in optical microscopy to break the diffraction-limit barrier, reliance of such techniques on fluorescent labeling technologies prohibits quantitative 3D imaging of the entire contents of cells. Cryoelectron microscopy can image pleomorphic structures at a resolution of 3—5 nm, but is only applicable to thin or sectioned speciments. Here, we report quantitative 3D imaging of a whole, unstained cell at a resolution of 50—60 nm by X-ray diffraction microscopy. We identified the 3D morphology and structure of cellular organelles including cell wall, vacuole, endoplasmic reticulum, mitochondria, granules, nucleus, and nucleolus inside a yeast spore cell. Furthermore, we observed a 3D structure protruding from the reconstructed yeast spore, suggesting the spore germination process. Using cryogenic technologies, a 3D resolution of 5—10 nm should be achievable by X-ray diffraction microscopy. This work hence paves a way for quantitative 3D imaging of a wide range of biological specimens at nanometer-scale resolutions that are too thick for electron microscopy.

• Publisher: National Academy of Sciences,National Acad Sciences
• Subject: Algorithms / Biological Sciences / cell nucleolus / cell walls / Cells / cryo-electron microscopy / Cryoelectron Microscopy - methods / Density / Diffraction patterns / Electron microscopes / Electrons / endoplasmic reticulum / fluorescent labeling / granules / image analysis / Image Processing, Computer-Assisted / Image reconstruction / Imaging / Imaging, Three-Dimensional - methods / Light microscopy / Microscopy / Mitochondria / Models, Biological / Models, Statistical / Optics and Photonics / Organelles / Physical Sciences / Pixels / Scattering, Radiation / Schizosaccharomyces - metabolism / Schizosaccharomyces - physiology / spore germination / Spores / Spores, Fungal - metabolism / Three dimensional imaging / vacuoles / X ray diffraction / X-Ray Diffraction - methods / X-Rays / Yeasts / Yeasts - metabolism