Cytoskeletal F-actin patterns quantitated with fluorescein isothiocyanate-phalloidin in normal and transformed cells

M. Verderame, D. Alcorta, M. Egnor, K. Smith, R. Pollack

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Abstract

Actin in cultured fibroblasts is organized into a complex set of fibers. Patterns of organization visualized with antibody to actin are similar but not identical to those visualized with fluorescein isothiocyanate-phalloidin (Fl-phalloidin), a chemical that binds to F-actin polymer with a dissociation constant of 2.7x10 -7M. Fl-phalloidin reveals that transformed cells have fewer, finer, and shorter F-actin-containing structures than do normal cells. Two-color fluorescence microscopy of single cells reveals that F-actin staining by Fl-phalloidin picks out the cytoskeletal cables more sharply than does antibody to actin, due to a reduced intracellular background fluorescence. This improved resolution permits sorting of cellular Fl-phalloidin patterns into four classes ranging in organization from 90% of the cytoplasm occupied by large cables to the absence of detectable cables. Reproducible differences in pattern distributions between normal and transformed cell lines have been quantitated. Fl-phalloidin together with rhodamine-based indirect antibody to simian virus 40 tumor antigen reveals a direct relationship between the degree of pattern change and simian virus 40 nuclear antigen expression in intermediate transformed 3T3 cell lines.

Original languageEnglish (US)
Pages (from-to)6624-6628
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume77
Issue number11 I
DOIs
Publication statusPublished - Dec 1 1980

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  • General

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