A 360° single‐axis tilt stage for the high‐voltage electron microscope

Abstract

A new type of specimen stage that permits more than 180° of tilting about the axis of a side-entry rod has been developed for a high-voltage electron microscope (HVEM). Roughly cylindrical specimens, with radial dimensions of less than a few micrometres, that can be mounted on the tip of a microneedle or micropipette are applicable. For glass micropipettes, the energy of the 1-MeV beam of the HVEM is sufficient to image specimens through both walls. The stage employs a spindle mechanism that holds these needles or micropipettes coaxial with the tilt axis, allowing the specimen to be rotated without restriction. This arrangement, along with the cylindrical form of the specimen, is an important development for single-axis tomography, because it permits a complete 180° set of projections to be recorded. The angular accuracy of the stage was demonstrated to be within ±0.20°, with a cumulative error of less than 1.0° over a 180° span. The new stage was tested using puffball spores mounted on a micropipette. A 180° tilt series was recorded and processed to yield a tomographic three-dimensional reconstruction which was displayed both as a cross-sectional view perpendicular to the tilt axis, and as a shaded surface viewed from different directions. The same computations were repeated using subsets of the tilt series to assess the effect of various amounts of missing information. Visual inspection of a selected cross-section from these reconstructions indicated that limiting the angular range to 160° produced results nearly as good as the full data set. Limiting the range to 140°, however, produced a noticeable geometric distortion, which became increasingly severe with ranges of 120° and 100°.