Open AccessSpecimen preparation for cryogenic coherent X‐ray diffraction imaging of biological cells and cellular organelles by using the X‐ray free‐electron laser at SACLA
Author(s) -
Kobayashi Amane,
Sekiguchi Yuki,
Oroguchi Tomotaka,
Okajima Koji,
Fukuda Asahi,
Oide Mao,
Yamamoto Masaki,
Nakasako Masayoshi
Publication year - 2016
Publication title -
journal of synchrotron radiation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.172
H-Index - 99
ISSN - 1600-5775
DOI - 10.1107/s1600577516007736
Subject(s) - diffraction , materials science , optics , laser , biological specimen , x ray crystallography , scanning electron microscope , electron microscope , free electron laser , composite material , physics
Coherent X‐ray diffraction imaging (CXDI) allows internal structures of biological cells and cellular organelles to be analyzed. CXDI experiments have been conducted at 66 K for frozen‐hydrated biological specimens at the SPring‐8 Angstrom Compact Free‐Electron Laser facility (SACLA). In these cryogenic CXDI experiments using X‐ray free‐electron laser (XFEL) pulses, specimen particles dispersed on thin membranes of specimen disks are transferred into the vacuum chamber of a diffraction apparatus. Because focused single XFEL pulses destroy specimen particles at the atomic level, diffraction patterns are collected through raster scanning the specimen disks to provide fresh specimen particles in the irradiation area. The efficiency of diffraction data collection in cryogenic experiments depends on the quality of the prepared specimens. Here, detailed procedures for preparing frozen‐hydrated biological specimens, particularly thin membranes and devices developed in our laboratory, are reported. In addition, the quality of the frozen‐hydrated specimens are evaluated by analyzing the characteristics of the collected diffraction patterns. Based on the experimental results, the internal structures of the frozen‐hydrated specimens and the future development for efficient diffraction data collection are discussed.