Application of a slow-scan CCD camera in protein electron crystallography at 400 KV

Abstract

In protein electron crystallography, high resolution amplitudes and phases are required for a 3D reconstruction of a protein in which one can trace the Cα-backbone. Recording of these amplitudes and phases has so far been done on photographic film. In principle they can be obtained with higher fidelity using an electronic device, such as a slow-scan charge-coupled device (CCD) camera. We use a 1024 × 1024 Gatan CCD camera (model 679) attached to a JEOL 4000EX intermediate voltage electron cryo-microscope. The camera has a 20-30 μm thin P43 phosphor scintillator and an extra set of fiber optics, that reduces the number of x-ray pixels in acquired frames. In this abstract, we will focus on several issues related to the CCD camera in obtaining high resolution diffraction patterns and images from thin protein crystals.To obtain electron diffraction patterns with sufficient statistical definition, one needs to use an adequate electron dose. However, at exposures well above 0.25 e/Å2, the pattern will suffer from blooming due to the central beam.