Three-dimensional electron microscopical visualization of the cytoskeleton of animal cells: immunoferritin identification of actin- and tubulin-containing structures

Abstract

Cytoskeletons prepared by Triton X-100 treatment of tissue culture cells appear in stereo electron microscopy as a highly organized and interconnected three-dimensional matrix of different fibrous elements. Microfilament bundles and also tonofilament-like bundles are readily discerned when present in the cell type. In addition thinner fibers, some of which branch (smallest diameter 30--40 A), as well as fibers of larger diameter, some of which correspond to microtubules, can be seen. Since such cytoskeletons are an open, membrane-free system, individual fibrous organizations can be identified by specific antibodies. An indirect immunoferritin procedure using antibodies to tubulin or actin visualizes microtubules or actin-containing structures. Stereo electron microscopy of cytoskeletons decorated with actin antibody reveals, in addition to the F-actin-containing microfilament bundles, an extended fine actin lattice. This actin net is displayed throughout the cytoplasm not only between the microfilament bundles but also in those regions of the cytoskeleton that in the intact cell correspond to the submembraneous regions. Thus all actin-containing fibrous cytoplasmic structures may be interconnected in the living cell.