Three-dimensional chromatin distribution in neuroblastoma nuclei shown by confocal scanning laser microscopy

Abstract

The relationship between cell shape and function has long been of interest. However, although the behaviour of the cytoskeleton during the cell cycle has been studied extensively variations in the shape and three-dimensional substructure of the nucleus are less well documented. The spatial distribution of chromatin has previously been studied by a mathematical analysis of the optical densities of stained nuclei, allowing an indirect derivation of the three-dimensional distribution of chromatin. More direct information on chromatin organization can be obtained from electron-microscopic serial sections, although this is very laborious. Using an iterative deconvolution algorithm, Agard and Sedat achieved a degree of optical sectioning in conventional fluorescence microscopy and reconstructed the three-dimensional arrangement of polytene chromosomes. We report here on the three-dimensional structure of cultured mammalian cells as visualized by confocal scanning laser microscopy (CSLM). The exceptionally short depth of field of this imaging technique provides direct optical sectioning which, together with its higher resolution, makes CSLM extremely useful for studying the three-dimensional morphology of biological structures.