ABSTRACT
Biological specimens are scarcely ideal for optical examination due to their structural heterogeneity, light absorption properties, refractive index and thickness. Such inherent attributes create serious problems in an investigation using any light microscope. Light and transmission electron microscopy (TEM) have traditionally been used to reveal the morphological changes associated with cellular development. Recently, the development of the confocal laser scanning microscope (CLSM) and its digital image acquisition system has made it possible to provide an alternative to other types of microscopy. In confocal microscopy both illumination and detection are confined to a single point in the specimen by inserting spiral filters (usually pin-holes) into the optical paths of the objective and condenser lenses. Confocal microscopy, with its various imaging modes and 3D-imaging capabilities, is particularly well-suited to detect the spatial organization of pollen by scanning at various pre-specified levels.
