Basic procedure of single-particle electron cryo-microscopy (CryoEM). The purified protein complex or assembly is frozen in a thin layer of vitrified ice (a) with presumably random orientation. The plus signs denote the positive static charges on the surface of ice illuminated by a high-energy electron beam, which is one of the reasons for image motion. The sample is then imaged to obtain different views (boxed particles in the panel (b) of the complex or assembly). Due to the low dose and corresponding low signal-to-noise-ratio (SNR) of the image, only the overall shape of particles is directly visible. After a 3D alignment procedure, many such particles are orientated and merged in 3D space to reconstruct a 3D density map (c). The side-chain details become visible in the map.