Alignment of electron tomographic series by correlation without the use of gold particles

Electron tomography requires that a tilt series of micrographs be aligned and that the orientation of the tilt axis be known. This has been done conveniently with gold markers applied to the surface of a specimen to provide easily accessible information on the orientation of each tilt projection. Where gold markers are absent, another approach to alignment must be used. A method is presented here for aligning tilt projections without the use of markers, utilizing correlation methods. The technique is iterative, drawing principally on the work of Dengler [Ultramicroscopy 30 (1989) 337], and consists of computing a low resolution back projection image from which computed tomographic projections can be generated. These in turn serve as reference images for the next alignment of the tomographic series. An initial alignment must be made before the first back projection, and this is done following the method of Guckenberger [Ultramicroscopy 9 (1982) 167] for translational alignment and by common lines analysis [Liu et al., Ultramicroscopy 58 (1995) 393] for identification of the tilt axis. Four tomographic series of a biological nature were aligned and analyzed, and the method has proven to be both accurate and reproducible for the data presented here.