Three-dimensional (3D) microstructure visualization on the electron microscope requires that the sample be tilted to different positions to collect a series of projections. This tilting should be performed rapidly for on-line stereo viewing and precisely for off-line tomographic reconstruction. Usually a projection series is collected using mechanical stage tilt alone. The stereo pairs must be viewed off-line and the 60 to 120 tomographic projections must be aligned with fiduciary markers or digital correlation methods. The delay in viewing stereo pairs and the alignment problems in tomographic reconstruction could be eliminated or improved by tilting the beam if such tilt could be accomplished without image translation.A microscope capable of beam tilt with simultaneous image shift to eliminate tilt-induced translation has been investigated for 3D imaging of thick (1 μm) biologic specimens. By tilting the beam above and through the specimen and bringing it back below the specimen, a brightfield image with a projection angle corresponding to the beam tilt angle can be recorded (Fig. 1a).
On-line observation of cell growth in a three-dimensional matrix on surface-modified microelectrode arrays
