Performing reflection-mode (backscatter-mode) confocal microscopy on
cells growing on reflective substrates gives images that have improved
contrast and are more easily interpreted than standard reflection-mode
confocal micrographs (Keith et al., 1998).
However, a number of factors degrade the quality of images taken with the
highest-resolution microscope objectives in this technique. We here
describe modifications to reflection-enhanced backscatter confocal
microscopy that (partially) overcome these factors. With these
modifications of the technique, it is possible to visualize structures the
size—and refractility—of individual microtubules in intact
cells. Additionally, we demonstrate that this technique, in common with
fluorescence techniques such as standing wave widefield fluorescence
microscopy and 4-Pi confocal microscopy, offers improved resolution in the
Z-direction.