Attachment of Tonofilaments to the Nuclear Membrane Demonstrated by HVEM Stereoscopy

Abstract

We have used thick section HVEM stereoscopy at multiple double-tilt angles to follow the course of long segments of tonofilament bundles in the squamous cells of human oral-cavity carcinoma and normal human skin. This approach serves to supplement and confirm the general distribution of the tonofilament network of whole cells indicated by light microscope immunofluorescent staining using specific keratin antibodies1. However, the HVEM approach has several advantages; (1) the drastic specimen preparation methods required in LM immunofluorescence to admit the antibody label into the cells can be avoided, (2) details of individual tonofilament attachment can be observed (3) the thick sections give better sampling of fine detail than can be obtained with conventional EM thin sections. Our initial thin section work gave no suggestion of attachment of tonofilaments to the nuclear membrane but this was frequently seen in the thick sections. On returning to the thin sections the probable attachments could be found. However, the absence of a 3-D view in the thin sections did not allow filament termination to be distinguished from grazing contact of filaments with the membrane. Finally, (4), the HVEM conveniently allows comparison of thick section results with critical-point-dried whole cells either prepared as for LM immunofluorescence or for HVEM.