TY - JOUR TI - Measuring the optimal exposure for single particle cryo-EM using a 2.6 Å reconstruction of rotavirus VP6 AU - Grant, Timothy AU - Grigorieff, Nikolaus A2 - Sundquist, Wesley I VL - 4 PY - 2015 DA - 2015/05/29 SP - e06980 C1 - eLife 2015;4:e06980 DO - 10.7554/eLife.06980 UR - https://doi.org/10.7554/eLife.06980 AB - Biological specimens suffer radiation damage when imaged in an electron microscope, ultimately limiting the attainable resolution. At a given resolution, an optimal exposure can be defined that maximizes the signal-to-noise ratio in the image. Using a 2.6 Å resolution single particle cryo-EM reconstruction of rotavirus VP6, determined from movies recorded with a total exposure of 100 electrons/Å2, we obtained accurate measurements of optimal exposure values over a wide range of resolutions. At low and intermediate resolutions, our measured values are considerably higher than obtained previously for crystalline specimens, indicating that both images and movies should be collected with higher exposures than are generally used. We demonstrate a method of using our optimal exposure values to filter movie frames, yielding images with improved contrast that lead to higher resolution reconstructions. This ‘high-exposure’ technique should benefit cryo-EM work on all types of samples, especially those of relatively low-molecular mass. KW - high-dose imaging KW - movie processing KW - radiation damage KW - optimal exposure KW - 20S proteasome KW - tomography JF - eLife SN - 2050-084X PB - eLife Sciences Publications, Ltd ER -