Electron microscopy and indirect immunofluorescence microscopy using monospecific tubulin antibodies were performed in parallel on glutaraldehyde-fixed tissue culture cells without osmium fixation. In order to reduce the excess aldehyde groups of the strongly crosslinked cellular matrix, which normally interfere with subsequent immunofluorescence microscopy, a mild NaBH4 treatment was introduced during or after the dehydration steps. Cells processed through the NaBH4 step show, in transmission electron microscopy, normal cytoplasmic microtubules ap roximately 250 A in diameter. When such cells are subjected to indirect immunofluorescence microscopy using monospecific tubulin antibody they reveal a complex system of unbroken, fine, fluorescent fibers traversing the cytoplasm between the perinuclear space and the plasma membrane. Thin sections of cells processed through the indirect immunofluorescence procedure show antibody-ecorated microtubules with a diameter of approximately 600 A. This decoration is not obtained when nonimmune IgGs are used instead of monospecific antitubulin IgGs. Thus, a direct comparison of cytoplasmic microtubules in glutaraldehyde-fixed cells by both electron microscopy and immunofluorescence microscopy can be obtained. Use of specific antibodies against actin and tubulin allows the distribution of microfilament bundles (1) and the organization of cytoplasmic microtubules (2) in tissue culture cells to be demonstrated by indirect immunofluorescence microscopy. In addition, the organization of tonofilaments has been demonstrated in one cell line by using an autoimmune serum (3). The advantages of the procedure include the direct overview provided over the whole cell and the opportunity to study numerous cells simultaneously.Documentation of cytoplasmic microtubules in tissue culture cells by immunofluorescence microscopy (2-10) has indicated the following features. (i) Microtubules are present in large numbers during interphase (2, 4). (ii) Microtubules can be followed for very long distances. They traverse the cytoplasm from the perinuclear space toward the plasma membrane and can also run for long distances underneath the plasma membrane (2, 4-10). (iii) Many of these cortical microtubules seem to originate in the centrosphere acting as an organization center, and after depolymerization they appear to regrow in a unidirectional manner toward the plasma membrane (5-7). (iv) At the onset of mitosis the complex pattern of cytoplasmic microtubules is reorganized to form the microtubules of the mitotic spindle. At late telophase, or in early G1, cytoplasmic microtubules reappear in the daughter cells originating from the centrosphere (4,(8)(9)(10).Although these results confirm and extend previous data on

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A special cell line derived from a rat mammary adenocarcinoma (RMCD cells) displays a distinct pattern of actomyosin fibrils (AM fibrils) visible with phase contrast, Nomarski interference and polarized light optics. It was shown that the cytoplasmic AM fibrils are arranged as bundles of highly parallel F-actin filaments. The chimical nature of the filaments was identified by incubation with heavy meromyosin from rabbit skeletal muscle. These cytoplasmic actomyosin fibrils actively contract under isotonic conditions. This was shown by contraction experiments under polarized light optics, by cinematographic analysis and by direct proof of the contractility of AM fibrils isolated by laser micro-dissection. Thus, cytoplasmic AM fibrils can be assumed to represent structures essential for motive force generation in contraction processes in non-muscle cells.