Published online by Cambridge University Press: 19 February 2014
In this study, a combined tilt- and focal series is proposed as a new recording scheme for high-angle annular dark-field scanning transmission electron microscopy (STEM) tomography. Three-dimensional (3D) data were acquired by mechanically tilting the specimen, and recording a through-focal series at each tilt direction. The sample was a whole-mount macrophage cell with embedded gold nanoparticles. The tilt–focal algebraic reconstruction technique (TF-ART) is introduced as a new algorithm to reconstruct tomograms from such combined tilt- and focal series. The feasibility of TF-ART was demonstrated by 3D reconstruction of the experimental 3D data. The results were compared with a conventional STEM tilt series of a similar sample. The combined tilt- and focal series led to smaller “missing wedge” artifacts, and a higher axial resolution than obtained for the STEM tilt series, thus improving on one of the main issues of tilt series-based electron tomography.
Current address: La Timone Hospital and Medicine School, Marseille, France
Combined tilt- and focal series high annular dark field scanning transmission electron microscopy (STEM) data. The sample was a whole mount macrophage cell containing gold nanoparticles of two different sizes distributed in clusters throughout its volume. The movie represents the aligned data recorded for a tilt range of -40° to +40° in 5° increments, with a focal series at each tilt angle using focus steps of 50 nm.
Perspective rendering of the reconstructed tomogram of the same cellular region as shown in Movie S1. The signal intensity was color-coded. The tilt-focal algebraic reconstruction technique (TF-ART) was used for the three dimensional reconstruction.