The Transmission Confocal Laser Scanning Microscope

Abstract A confocal laser scanning microscope which can collect images in both transmission and reflection modes has been installed and is being tested in the Imaging Laboratories of the John P. Roberts Research Institute, London, Ontario. Designed by Dr. Ted Dixon at the University of Waterloo, it is being developed by a multi-disciplinary research group which includes the Ontario Lasers and Lightwaves Research Centre and the Department of Physiology, University of Toronto; the Radiology, Physics, and Pathology Departments, MacMaster University; and the Zoology Department, University of Western Ontario. Commercial confacal microscopes operate by reflectance or epifluorescence. A pair of scanning mirrors direct a diffuse laser beam in a raster pattern through an objective lens, which focuses it on the specimen. Reflected light passes back along the same light path, being “de-scanned” by the moving mirrors and then diverted by a beam splitter into a detector. On its return to the detector, light from the focal plane is focussed through a pinhole, which blocks light from out-of-focus regions of the spectrum The microscope stage is moved up and down by a stepping motor to collect images at different depths.