Zero‐loss energy filtering under low‐dose conditions using a post‐column energy filter

Electron cryomicroscopy combined with energy filtering can be performed under low‐dose conditions using a post‐column energy filter. The microscope combined with the filter is set up such that it can be used with similar ease as a conventional microscope, the main difference being that all filter and microscope control is performed through a central computer and images are recorded with a cooled slow‐scan CCD camera. The microscope can also still be used for regular imaging on film as without the filter. Owing to the 18 times post‐magnification of the filter, the microscope normally has to be operated at a small magnification, e.g. 3000×, and the beam has to be contracted to a small spot, e.g. 5 mm, in the plane of the microscope viewing screen. Computer control allows one to perform a variety of tasks automatically, such as autofocusing, thickness measurements, most‐probable‐loss imaging, CCD spot‐scanning and tomography. The gain in contrast due to zero‐loss energy filtering is analysed using visual inspection, power spectra and Fourier ring correlation. The thickness range for ice‐embedded specimens in which a filter at 120 kV is most useful appears to be between 100 and 300 nm.