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SUMMARY When the zero‐loss transmission falls below 10−3 for biological sections of mass‐thickness greater than 70 μg/cm2, the energy window in the electron spectroscopic imaging (ESI) mode of an energy‐filtering electron microscope (EFEM) can be shifted to the most‐probable energy loss of the electron energy‐loss spectrum. This enables mass‐thicknesses up to 150 μg/cm2 or thicknesses of 1·5 μm to be examined. Electron energy‐loss spectra of thick carbon films calculated by a Fourier method agree with experimental spectra. Measurements of the electron energy‐loss spectroscopy and ESI image intensities with an additional platinum film confirm a scattering model for the calculation of the image intensity. This model considers the angular broadening at the most‐probable energy loss by introducing an effective illumination aperture of the order of the full‐width at half‐maximum of the angular distribution.

SUMMARY An energy‐filtering microscope working at 80 keV is used for the investigation of the effect of inelastic scattering on Bragg contrast. Inelastic scattering results in a preservation of Bragg contrast but edge and bend contours are blurred by a spectrum of excitation errors due to the angular distribution of inelastic scattering. This blurring and the chromatic aberration results in a decrease of contrast and resolution for thick specimens. Therefore, contrast and resolution can be increased by zero‐loss filtering as shown by evaporated films of increasing thickness below 150 μg/cm2. Up to ∼300 μg/cm2 an energy‐filtered image at the most probable energy shows the best results. The results obtained are extrapolated to energy‐filtered high‐voltage electron microscopy.