A procedure was developed for the conjugation of vimentin with biotin. Biotinylated vimentin was then microinjected into BHK-21 cells and the fate of the labeled protein was determined at various times postinjection by indirect immunofluorescence. Microinjected vimentin could be traced through a specific sequence of morphological changes ultimately resulting in the formation of a filamentous network. The injected protein was first detected in spots dispersed throughout the cytoplasm. Subsequently, these spots appeared to cluster near the nucleus where they merged into a diffuse 'cap'. This cap coincided with a concentration of endogenous intermediate filaments and eventually gave rise to a filamentous network that was coincident with the endogenous intermediate filament network as a determined by double-label immunofluorescence. The results indicate that the incorporation of exogenous vimentin into a filamentous network is initiated in a perinuclear region and progresses in a polarized fashion toward the cell surface.
Vikstrom, K. L., Borisy, G. G., & Goldman, R. D. (1989). Dynamic aspects of intermediate filament networks in BHK-21 cells. Proceedings of the National Academy of Sciences of the United States of America, 86(2), 549–553. https://doi.org/10.1073/pnas.86.2.549