Image reconstruction using a focus series on thick biological specimens: TEM CTF correction

Abstract

The primary project of our laboratory is the investigation of chromatin structure by three dimensional electron microscope tomography. Three dimensional tomography involves the reconstruction of an object by combining multiple projection views of the object at different tilt angles. There are two complications that give rise to an inaccurate representation in the image of the projected object mass density: 1) electron-specimen interactions, and 2) electron microscope lens aberrations. In our abstract, “Analysis of Electron-specimen Interactions of Thick Biological Specimens in Transmission Electron Microscopy at 200 keV” elsewhere in these proceedings, we investigated image formation at the level of electron-specimen interactions at 200 keV. We extrapolated that at 300 keV, there is still a significant amount of elastic scattering for thick (up to 0.7 microns) biological specimens . Here, we demonstrate that this allows the correction of the microscope aberration function through multiple focus observations can enhance contrast.For high resolution electron microscopy of crystalline materials, some authors have demonstrated that the focus variation method to reconstruct the specimen exit-surface electron wave can lead to an improvement in resolution.