High resolution structural analysis of macromolecular assemblies and cellular architectures is essential for understanding the mechanisms of biological functions because they are all determined by the structures and dynamics of the molecules and cells at atomic level. Although X-ray crystallography and NMR are powerful tools for structural analysis of macromolecules, the difficulty in crystallization and the size limit of target molecules pause severe limitations on the application of the former and the latter, respectively. Filamentous protein complexes are particularly difficult target because the length is not uniform. Electron cryomicroscopy (cryoEM) and image analysis is a powerful tool because there is no size limitation and it requires only a very small amount of sample for data collection. The achievable resolution, however, tends to be limited by intrinsically low signal to noise ratio (S/N) of cryoEM images due to extremely low electron dose to avoid radiation damage. Therefore, tens of thousands of particle images have to be collected, classified according to the orientation, aligned and averaged to reconstruct high-resolution 3D images. However, accurate image alignment is difficult due to the intrinsically low S/N of cryoEM images. We developed techniques to improve the image S/N and speed of image collection by the use of an energy filter and CCD camera in combination with the liquid helium-cooled specimen stage on an electron microscope with a field emission electron gun operated at 200–300 kV. We have gained nearly 5 times higher S/N than before by controlling ice thickness and raising the specimen temperature from 4 K to around 50 K. The undesirable charging effect has also been minimized at 50 K, allowing us to use nearly all the collected images for analysis, while only a few % of images were usable at 4 K. Single particle image analysis now allows the high-resolution structural analysis of filamentous protein complexes to be completed within a week, and the achievable resolution is beyond 4 Å. I will describe the structures of various filamentous protein complexes and discuss the potential of the method for future life sciences.