A new cryo-EM system for single particle analysis.

• A cryo-EM with a cold-field emission gun (CFEG) was tested for single particle analysis. • GUI programs were developed for an efficient and accurate data collection. • Imaging with CFEG is beneficial to obtaining high-resolution structures by single-particle cryo-EM. A new cryo-EM system has been investigated for single particle analysis of protein structures. The system provides parallel illumination of a highly-coherent 300 kV electron beam from a cold-field emission gun, and boosts image contrast with an in-column energy filter and a hole-free phase plate. It includes motorized cryo-sample loading and automated liquid-nitrogen filling for cooling multiple samples. In this study, we describe gun and electron beam characteristics, and demonstrate the suitability of this system for single particle reconstructions. The performance of the system is tested on two examples, a spherical virus and apoferritin. GUI programs have also been developed to control and monitor the system for correct illumination, imaging with less ellipticity and steady magnification, and timing of flashing and liquid-nitrogen filling. These programs are especially useful for efficient application of the system to single particle cryo-EM. [ABSTRACT FROM AUTHOR]