High voltage microscopy of biological specimens: Some practical considerations

SUMMARY Techniques are now available for the preparation of specimens giving excellent image contrast at high voltages, either by block‐staining, staining an already embedded block or by post‐staining. Further techniques are being developed for selectively staining certain features of the specimen thereby reducing the stain content of the specimen and permitting the examination of very thick specimens at adequate resolution. Specimens having inadequate contrast, particularly preparations of macro‐molecules, may be imaged in one of the various dark‐field modes, giving a considerable enhancement in image contrast on the viewing screen. Further enhancement of the recorded image contrast can be achieved by exposing the photographic plate to a much higher density than is normal. The complexity of image detail which occurs when the many features of a thick specimen are contracted into a two‐dimensional micrograph may be elucidated by stereo‐imaging techniques. Care must be taken to optimize the stereo‐tilt angles, the orientation of the tilt axis and the register of the two micrographs in the stereo‐pair, for the maximum depth in the specimen to be observed.