表征阿波罗直接电子探测器的分辨率和吞吐量

IF 3.5 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY Journal of Structural Biology: X Pub Date : 2023-01-01 DOI:10.1016/j.yjsbx.2022.100080
Ruizhi Peng , Xiaofeng Fu , Joshua H. Mendez , Peter S. Randolph , Benjamin E. Bammes , Scott M. Stagg
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引用次数: 4

摘要

电子探测技术的进步对高分辨率低温电镜结构测定的成功至关重要。新一代的直接电子探测器被称为阿波罗,已经由直接电子公司开发。阿波罗使用了一种新颖的基于事件的map探测器,专为超快速电子计数而设计。我们对这款新相机进行了评估,发现它提供了高探测量子效率(DQE)和低重合损耗,能够以每秒每像素近80个输入电子的速度获得高质量的电子计数数据。我们进一步表征了Apollo在真实生物样品上的单粒子低温电镜性能。使用小鼠载铁蛋白,阿波罗在所有三种测试剂量率下获得了优于1.9 Å的分辨率重建。随着采集时间的延长和标本制备方法的改进,小鼠载铁蛋白重构至1.66 Å分辨率。应用于更具挑战性的小蛋白醛缩酶,我们获得了2.24 Å分辨率重建。高质量的地图表明阿波罗具有足够高的DQE,可以高保真地重建较小的蛋白质和复合物。我们的研究结果表明,阿波罗相机在广泛的剂量率范围内表现良好,能够捕获高质量的数据,为大小单颗粒样品产生高分辨率的重建。
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Characterizing the resolution and throughput of the Apollo direct electron detector

Advances in electron detection have been essential to the success of high-resolution cryo-EM structure determination. A new generation of direct electron detector called the Apollo, has been developed by Direct Electron. The Apollo uses a novel event-based MAPS detector custom designed for ultra-fast electron counting. We have evaluated this new camera, finding that it delivers high detective quantum efficiency (DQE) and low coincidence loss, enabling high-quality electron counting data acquisition at up to nearly 80 input electrons per pixel per second. We further characterized the performance of Apollo for single particle cryo-EM on real biological samples. Using mouse apoferritin, Apollo yielded better than 1.9 Å resolution reconstructions at all three tested dose rates from a half-day data collection session each. With longer collection time and improved specimen preparation, mouse apoferritin was reconstructed to 1.66 Å resolution. Applied to a more challenging small protein aldolase, we obtained a 2.24 Å resolution reconstruction. The high quality of the map indicates that the Apollo has sufficiently high DQE to reconstruct smaller proteins and complexes with high-fidelity. Our results demonstrate that the Apollo camera performs well across a broad range of dose rates and is capable of capturing high quality data that produce high-resolution reconstructions for large and small single particle samples.

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来源期刊
Journal of Structural Biology: X
Journal of Structural Biology: X Biochemistry, Genetics and Molecular Biology-Structural Biology
CiteScore
6.50
自引率
0.00%
发文量
20
审稿时长
62 days
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