High-resolution analysis of the 3D organization of human metaphase chromosomes.

The detailed understanding of the nuclear cell functions requires an accurate knowledge of the spatial organization of their structures. For many years the study of human metaphase chromosomes was conducted with light microscopy after staining protocols disturbing the native chromosomal structure. The approach by scanning electron microscopy (SEM) provides higher resolution as compared with light microscopy and permits surface analysis of the chromosomal structure, which cannot be adequately obtained from transmission electron microscopy. Nevertheless, to obtain high resolution in SEM observations, the use of a high electron accelerating voltage (up to 30 kV) is required (1). With these experimental conditions, the sputter coating or a conductive staining of the samples is generally required (1,2). Both procedures allow electron-charging dispersion from the sample but may obscure fine details and produce sample alterations (3).