Performance of a slow-scan CCD camera for macromolecular imaging in a 400 kV electron cryomicroscope

The feasibility and limitations of a 1024 x 1024 slow-scan charge-coupled device (CCD) camera were evaluated for imaging in a 400 kV electron cryomicroscope. Catalase crystals and amorphous carbon film were used as test specimens. Using catalase crystals, it was found that the finite (24 μm) pixel size of the slow-scan CCD camera governs the ultimate resolution in the acquired images. For instance, spot-scan images of ice-embedded catalase crystals showed resolutions of 8 Å and 4 Å at effective magnifications of 67,000 x and 132,000 x, respectively. Using an amorphous carbon film, the damping effect of the modulation transfer function (MTF) of the slow-san CCD camera on the specimen's Fourier spectrum relative to that of the photographic film was evaluated. The MTF of the slow-san CCD camera fell off more rapidly compared to that of the photographic film and reached the value of 0.2 at the Nyquist frequency. Despite this attenuation, the signal-to-noise ratio of the CCD data, as determined from reflections of negatively-stained catalase crystals, was found to decrease to ~50% of that of photographic film data. The phases computed from images of the same negatively-stained catalase crystals recorded consecutively on both the slow-scan CCD camera and photographic film were found to be comparable to each other within 12°. Ways of minimizing the effect of the MTF of the slow-scan CCD camera on the acquired images are also presented.