An algorithm for estimation and correction of anisotropic magnification distortion of cryo-EM images without need of pre-calibration.
An algorithm for estimation and correction of anisotropic magnification distortion of cryo-EM images without need of pre-calibration.
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DOI：
10.1016/j.jsb.2016.06.003
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发表时间：
2016-08
影响因子：
3
通讯作者：
Jiang W
中科院分区：
文献类型：
--
作者：
Yu G;Li K;Liu Y;Chen Z;Wang Z;Yan R;Klose T;Tang L;Jiang W
在单粒子三维重建中，电子显微镜图像的各向异性放大失真(主要是椭圆失真)是一个潜在的分辨率限制因素。据报道，在低放大倍率(如18000x)下，多台显微镜的椭圆失真约为1-3%，这大大限制了单粒子三维重建的分辨率，特别是对大粒子。在这里，我们报告了一种通用算法，它将失真校正问题描述为一个广义的二维对准任务，并直接从粒子图像估计失真参数。与目前的预校准方法不同，我们的计算方法适用于使用任何显微镜在广泛的放大范围内收集的所有数据集，而不需要额外的实验测量。此外，我们的方法中的每显微图像和/或每粒子级别的椭圆失真估计可以解决低温EM数据集中潜在的失真变化，并通过预校准方法进一步改善相对于常值校正的三维重建。通过对多个数据集的成功应用和预校准方法的交叉验证，我们证明了该算法在估计失真方面的有效性和健壮性；椭圆失真的校正显著提高了可实现的分辨率~1-3倍，并能够在2.4-2.6？分辨率下对多个病毒结构进行三维重建。研究发现，具有椭圆失真的分辨率极限和经过失真校正的分辨率改进量与粒子大小和畸变量的乘积密切相关，这有助于评估椭圆失真是否是单粒子低温电磁项目的主要分辨率限制因素。
Anisotropic magnification distortion of TEM images (mainly the elliptic distortion) has been recently found as a potential resolution-limiting factor in single particle 3-D reconstruction. Elliptic distortions of ~1–3% have been reported for multiple microscopes under low magnification settings (e.g., 18,000x), which significantly limited the achievable resolution of single particle 3-D reconstruction, especially for large particles. Here we report a generic algorithm that formulates the distortion correction problem as a generalized 2-D alignment task and estimates the distortion parameters directly from the particle images. Unlike the present pre-calibration methods, our computational method is applicable to all datasets collected at a broad range of magnifications using any microscope without need of additional experimental measurements. Moreover, the per-micrograph and/or per-particle level elliptic distortion estimation in our method could resolve potential distortion variations within a cryo-EM dataset, and further improve the 3-D reconstructions relative to constant-value correction by the pre-calibration methods. With successful applications to multiple datasets and cross-validation with the pre-calibration method, we have demonstrated the validity and robustness of our algorithm in estimating the distortion; Correction of the elliptic distortion significantly improved the achievable resolutions by ~1–3 folds and enabled 3-D reconstructions of multiple viral structures at 2.4–2.6 Å resolutions. The resolution limits with elliptic distortion and the amounts of resolution improvements with distortion correction were found to strongly correlate with the product of the particle size and the amount of distortion, which can help assess if elliptic distortion is a major resolution limiting factor for single particle cryo-EM projects.