ジャーナル: J Struct Biol / 年: 2013 タイトル: Maximizing the potential of electron cryomicroscopy data collected using direct detectors. 著者: David Veesler / Melody G Campbell / Anchi Cheng / Chi-Yu Fu / Zachary Murez / John E Johnson / Clinton S Potter / Bridget Carragher / 要旨: Single-particle electron cryomicroscopy is undergoing a technical revolution due to the recent developments of direct detectors. These new recording devices detect electrons directly (i.e. without ...Single-particle electron cryomicroscopy is undergoing a technical revolution due to the recent developments of direct detectors. These new recording devices detect electrons directly (i.e. without conversion into light) and feature significantly improved detective quantum efficiencies and readout rates as compared to photographic films or CCDs. We evaluated here the potential of one such detector (Gatan K2 Summit) to enable the achievement of near-atomic resolution reconstructions of biological specimens when coupled to a widely used, mid-range transmission electron microscope (FEI TF20 Twin). Compensating for beam-induced motion and stage drift provided a 4.4Å resolution map of Sulfolobus turreted icosahedral virus (STIV), which we used as a test particle in this study. Several motion correction and dose fractionation procedures were explored and we describe their influence on the resolution of the final reconstruction. We also compared the quality of this data to that collected with a FEI Titan Krios microscope equipped with a Falcon I direct detector, which provides a benchmark for data collected using a high-end electron microscope.
凍結剤: ETHANE / チャンバー内温度: 94 K / 装置: GATAN CRYOPLUNGE 3 / 詳細: Vitrification was carried out at room temperature. / 手法: Blot for 3 seconds before plunging.
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電子顕微鏡法
顕微鏡
FEI TECNAI F20
温度
平均: 90 K
アライメント法
Legacy - 非点収差: Objective lens astigmatism was corrected at 41322 times magnification
日付
2012年11月30日
撮影
カテゴリ: CCD / フィルム・検出器のモデル: GATAN K2 (4k x 4k) / 実像数: 754 / 平均電子線量: 22 e/Å2 詳細: Every movie is composed of sixteen frames recorded by the direct electron detector.
The final reconstruction was sharpened with a negative temperature factor of 650 A^2.
CTF補正
詳細: Each particle
最終 再構成
アルゴリズム: OTHER / 解像度のタイプ: BY AUTHOR / 解像度: 5.8 Å / 解像度の算出法: OTHER / ソフトウェア - 名称: Frealign 詳細: The final reconstruction was computed using the first ten frames of each movie and sharpened with a negative temperature factor of 650 A^2. 使用した粒子像数: 4446