Conditions for electron tomographic data acquisition.

The conventional algorithms employed in electron microscope tomography require that the series of images obtained from different orientations of the specimen each represent a parallel orthographic projection under uniform magnification. Electron microscope optics can produce distortions in images that may affect the accuracy of a tomographic reconstruction. These distortions result in images with differential rotation and magnification of regions of a thick or highly tilted specimen located at different distances with respect to the plane of focus. The distortions increase in magnitude and may significantly affect the accuracy of the tomographic data when images are acquired under current center misalignment or non-eucentric positioning of the specimen. An additional source of error can be introduced if the condenser lenses are adjusted to compensate for the intensity attenuation due to the increased beam path length as the specimen is tilted. The change in beam crossover position due to adjustment of the condensers alters the beam divergence and may introduce systematic changes in magnification over the tilt series. The process of alignment of the tilt series will propagate these errors to more central regions of the image. In most cases, with proper consideration, it is possible to minimize these errors to levels where they will have negligible effects on the resolution of the tomographic reconstruction.