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利用冷冻软 X 射线断层摄影术对细胞进行三维绘图描述。

A 3D Cartographic Description of the Cell by Cryo Soft X-ray Tomography.

机构信息

MISTRAL beamline, Alba Light Source, Cerdanyola del Valles, Spain.

Beamline B24, Diamond Light Source, Harwell Science and Innovation Campus, Didcot, UK.

出版信息

J Vis Exp. 2021 Mar 15(169). doi: 10.3791/62190.

DOI:10.3791/62190
PMID:33779598
Abstract

Imaging techniques are fundamental in order to understand cell organization and machinery in biological research and the related fields. Among these techniques, cryo soft X-ray tomography (SXT) allows imaging whole cryo-preserved cells in the water window X-ray energy range (284-543 eV), in which carbon structures have intrinsically higher absorption than water, allowing the 3D reconstruction of the linear absorption coefficient of the material contained in each voxel. Quantitative structural information at the level of whole cells up to 10 µm thick is then achievable this way, with high throughput and spatial resolution down to 25-30 nm half-pitch. Cryo-SXT has proven itself relevant to current biomedical research, providing 3D information on cellular infection processes (virus, bacteria, or parasites), morphological changes due to diseases (such as recessive genetic diseases) and helping us understand drug action at the cellular level, or locating specific structures in the 3D cellular environment. In addition, by taking advantage of the tunable wavelength at synchrotron facilities, spectro-microscopy or its 3D counterpart, spectro-tomography, can also be used to image and quantify specific elements in the cell, such as calcium in biomineralization processes. Cryo-SXT provides complementary information to other biological imaging techniques such as electron microscopy, X-ray fluorescence or visible light fluorescence, and is generally used as a partner method for 2D or 3D correlative imaging at cryogenic conditions in order to link function, location, and morphology.

摘要

成像技术对于理解生物学研究和相关领域中的细胞组织和机制至关重要。在这些技术中,冷冻软 X 射线断层扫描(Cryo-SXT)允许在水窗 X 射线能量范围内(284-543 eV)对整个冷冻保存的细胞进行成像,在这个范围内,碳结构的固有吸收率高于水,从而可以对每个体素中包含的材料的线性吸收系数进行 3D 重建。通过这种方式,可以实现高达 10 µm 厚的整个细胞的定量结构信息,具有高通量和空间分辨率,低至 25-30nm 半节距。Cryo-SXT 已被证明与当前的生物医学研究相关,提供有关细胞感染过程(病毒、细菌或寄生虫)、由于疾病(如隐性遗传疾病)引起的形态变化的 3D 信息,并帮助我们了解药物在细胞水平上的作用,或在 3D 细胞环境中定位特定结构。此外,通过利用同步加速器设施的可调谐波长,可以使用光谱显微镜或其 3D 对应物,即光谱断层扫描,对细胞中的特定元素进行成像和定量,例如生物矿化过程中的钙。Cryo-SXT 为电子显微镜、X 射线荧光或可见光荧光等其他生物成像技术提供了互补信息,通常作为低温条件下 2D 或 3D 相关成像的辅助方法,以将功能、位置和形态联系起来。

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