Imaging modes for bilateral confocal scanning microscopy

Abstract

SummaryThe bilateral scanning approach to confocal microscopy is characterized by the direct generation of the image on a two‐dimensional (2‐D) detector. This detector can be a photographic plate, a CCD detector or the human eye, the human eye permitting direct visualization of the confocal image. Unlike Nipkow‐type systems, laser light sources can be used for excitation. A design called a carousel has been developed, in which the bilateral confocal scan capability can be added to an existing microscope so that rapid exchange and comparison between confocal and non‐confocal imaging conditions is possible. The design permits independent adjustment of confocal sectioning properties with lateral resolutions better than, or, in the worst case equivalent to, those available in conventional microscopy. The carousel can be considered as a stationary optical path in which certain imaging conditions, such as confocality, are defined and operate on part of the imaging field. The action of the bilateral scan mirror then extends this image condition over the whole field. A number of optical arrangements for the carousel are presented which realize various forms of confocal fluorescence and reflection imaging, with point, multiple point or slit confocal detection arrangements. Through the addition of active elements to the carousel direct stereoscopic, ratio, time‐resolved and other types of imaging can be achieved, with direct image formation on a CCD, eye or other 2‐D detectors without the need to modify the host microscope. Depending on the photon flux available, these imaging modes can run in real‐time or can use a cooled CCD at (very) low light level for image integration over an extended period.