A scanning electron microscopic technique for three-dimensional visualization of the spatial arrangement of...

Chromosome and chromatid alignment in mitotic configurations remains a topic of interest because there is little precise information. For example, reconstruction of mitotic configurations from serial sections collected with transmission electron microscopy has proven to be neither practical nor a sensitive method for conceptualizing these arrangements. Similarly light microscopy has been even more unsatisfactory because of its limited resolution and lack of three-dimensional capabilities. These limitations conceivably could be overcome by visualization of mitotic configurations by scanning electron microscopy (SEM). However, SEM has its limitations, of which the most obvious with regard to visualization of mitotic configurations, is that such structures in dividing cells are obscured from the beam by membranes, cellular organelles, and the mitotic apparatus. These "contaminants," we have found, can be removed by the appropriate procedure such that a direct three-dimensional visualization of intact life-like mitotic configurations of chromatids from mammalian cells is possible. We also demonstrate that these configurations, although some artifacts may exist, retain the same basic shape and chromatid arrangements throughout metaphase, anaphase, and telophase when compared to configurations isolated with a non-ionic detergent and neutral buffers.