The image obtained in a transmission electron microscope is the two-dimensional projection of a three-dimensional (3D) object. The 3D reconstruction of the object can be calculated from a series of projections by back-projection, but this algorithm assumes that the image is linearly related to a line integral of the object function. However, there are two kinds of contrast in electron microscopy, scattering and phase contrast, of which only the latter is linear with the optical density (OD) in the micrograph. Therefore the OD can be used as a measure of the projection only for thin specimens where phase contrast dominates the image. For thick specimens, where scattering contrast predominates, an exponential absorption law holds, and a logarithm of OD must be used. However, for large thicknesses, the simple exponential law might break down due to multiple and inelastic scattering.
Angular range, sampling and noise considerations for inverse light scattering analysis of nuclear morphology