2D-crystallization in combination with transmission electron microscopy (TEM) is one of the few methods for the structural analysis of membrane proteins in their native state. However, the parameters for the generation of large crystalline sheets are typically difficult to identify for a given protein. Many repetitive and time consuming screening steps by TEM are therefore necessary to find the best crystallization and preparation conditions. Although several software packages offer the possibility to control an electron microscope, none is completely adapted for a fully automated and completely integrated acquisition and analysis of 2D crystals. Here we report on the development of a fully automatic screening and on-line analysis software for the fast and automatic survey of large quantities of negatively stained EM samples for 2D crystallography.