IceBreaker: Software for high-resolution single-particle cryo-EM with non-uniform ice.

Despite the abundance of available software tools, optimal particle selection is still a vital issue in single-particle cryoelectron microscopy (cryo-EM). Regardless of the method used, most pickers struggle when ice thickness varies on a micrograph. IceBreaker allows users to estimate the relative ice gradient and flatten it by equalizing the local contrast. It allows the differentiation of particles from the background and improves overall particle picking performance. Furthermore, we introduce an additional parameter corresponding to local ice thickness for each particle. Particles with a defined ice thickness can be grouped and filtered based on this parameter during processing. These functionalities are especially valuable for on-the-fly processing to automatically pick as many particles as possible from each micrograph and to select optimal regions for data collection. Finally, estimated ice gradient distributions can be stored separately and used to inspect the quality of prepared samples.