Electron beam damage is the fundamental limit to resolution in electron cryomicroscopy (cryo-EM) of frozen, hydrated specimens. Radiation damage increases with the number of electrons used to obtain an image and affects information at higher spatial frequencies before low-resolution information. For the experimentalist, a balance exists between electron exposures sufficient to obtain a useful signal-to-noise ratio (SNR) in images and exposures that limit the damage to structural features. In single particle cryo-EM this balance is particularly delicate: low-resolution features must be imaged with a sufficient SNR to allow image alignment so that high-resolution features recorded below the noise level can be recovered by averaging independent images. By measuring the fading of Fourier components from images obtained at 200 kV of thin crystals of catalase embedded in ice, we have determined the electron exposures that will maximize the SNR at resolutions between 86 and 2.9 angstrom. These data allow for a rational choice of exposure for single particle cryo-EM. For example, for 20 angstrom resolution, the SNR is maximized at similar to 20 e(-)/angstrom(2), whereas for 3 angstrom resolution, it is maximized at similar to 10 e(-)/angstrom(2). We illustrate the effects of exposure in single particle cryo-EM with data collected at similar to 12-15 and similar to 24-30 e(-)/angstrom(2). (C) 2009 Elsevier Inc. All rights reserved.