The presence of preferred orientations in single particle analysis (SPA) by cryo-Electron Microscopy (cryoEM) is currently one of the hurdles preventing many structural analyses from yielding high-resolution structures. Although the existence of preferred orientations is mostly related to the grid preparation, in this technical note, we show that some image processing algorithms used for angular assignment and three-dimensional (3D) reconstruction are more robust than others to these detrimental conditions. We exemplify this argument with three different data sets in which the presence of preferred orientations hindered achieving a 3D reconstruction without artifacts or, even worse, a 3D reconstruction could never be achieved.

通过低温电子显微镜 (cryoEM) 进行的单粒子分析 (SPA) 中优选取向的存在是目前阻止许多结构分析产生高分辨率结构的障碍之一。尽管首选方向的存在主要与网格准备有关，但在本技术说明中，我们展示了一些用于角度分配和三维 (3D) 重建的图像处理算法对这些不利条件比其他算法更稳健。我们用三个不同的数据集举例说明了这一论点，其中首选方向的存在阻碍了无伪影的 3D 重建，或者更糟糕的是，永远无法实现 3D 重建。