Fine structures of human chromosomes observed by X-ray contact microscopy coupled with atomic force microscopy

Chromosomes from human lymphocytes were observed by X-ray contact microscopy (XRCM) using laser-produced plasma X-rays coupled with atomic force microscopy (AFM). A resist with either dry or hydrated chromosomes was exposed to laser-produce, plasma x-rays. The exposed resist was then developed after removing all cell-derived materials including chromosomes and was observed directly by AFM. Clear images of fine structures of chromosomes in various periods of the cell cycle could be obtained. It was shown that AFM is a powerful tool for XRCM for its easiness to locate the X-ray images on a resist once pinpointed by light microscopy in comparison with transmission electron microscopy (TEM) in combination with replica method. In addition, the data suggested that chromatin fiber is composed basically of linearly arranged particles with heterogeneous sizes, which is incompatible with the current textbook model regarding the conformation of chromatin where a nucleosomal fiber coils to generate a chromatin fiber with the homogeneous size of ca. 30 nm in diameter.