Visualization of cytoskeletal elements in the transmission X-ray microscope

Transmission X-ray microscopy has been used to study the arrangement of cytoskeletal filaments in interphase PtK2 cells. Extraction of the soluble proteins and of some organelles with the nonionic detergent Triton X-100 was important in obtaining sufficient image contrast between the insoluble cytoskeletal filaments and the surrounding cytoplasm. If this step is not performed cytoskeletal filaments are not visualized and transmission X-ray micrographs of the cytoplasm instead show predominantly membrane-bound organelles such as vesicles and the endoplasmic reticulum. Transmission X-ray micrographs of the cytoskeletal filaments and endoplasmic reticulum in air-dried specimens, as well as in specimens examined in the wet state, can be directly compared with transmission electron micrographs of cytoskeletons prepared in the same way. The profiles seen with the two techniques are similar, although, currently, transmission X-ray micrographs have a limit of resolution of similar to 50 nm. Transmission X-ray micrographs appear to show some substructure in interphase nuclei in cells fixed either with glutaraldehyde or by cryofixation and examined in a hydrated condition. (C) 1998 Academic Press.