Abstract
Cytoplasmic microtubules are noncovalent polymers of the protein tubulin. In the cells, the main function of microtubules is to provide tracks for organelle transport. Two experimental approaches based on fluorescence microscopy are commonly used to examine organization of microtubules in mammalian tissue culture cells. The first experimental approach involves indirect immunofluorescence staining of chemically fixed cells with tubulin antibody. Fluorescence microscopy of immunostained specimens allows the examination of the distribution of microtubules in the cytoplasm at the moment of fixation. The second experimental approach involves introduction of tubulin subunits covalently labeled with a fluorochrome into the cytoplasm of living cells. Time-lapse fluorescence microscopy of cells containing labeled tubulin subunits allows to examine changes in the spatial organization of microtubules in the cytoplasm and also to directly observe their behavior. In this chapter, we describe preparation of samples for fluorescence microscopy of microtubules.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Desai, A. and Mitchison, T. J. (1997) Microtubule polymerization dynamics. Ann. Rev. Cell Dev. Biol. 13, 83ā117.
Komarova, J., Peloquin, J., and Borisy, G. (2004) Microinjection of fluorophore-labeled proteins, in Live Cell Imaging, (Goldman, R. D. and Spector, D. L., ed.), Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, pp. 67ā86.
Goodson, H. V. and Wadsworth, P. (2004) Methods for expressing and analyzing GFP-tubulin and GFP-microtubule-associated proteins, in Live Cell Imaging, (Goldman, R. D. and Spector, D. L., ed.), Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, pp. 537ā553.
Rusan, N. M., Fagerstrom, C. J., Yvon, A. M. C., and Wadsworth, P. (2001) Cell cycle-dependent changes in microtubule dynamics in living cells expressing green fluorescent protein-Ī± tubulin. Mol. Biol. Cell 12, 971ā980.
Borisy, G. G., Marcum, J. M., Olmsted, J. B., Murphy, D. B., and Johnson, K. A. (1975) Purification of tubulin and associated high molecular weight proteins from porcine brain and characterization of microtubule assembly in vitro. Ann. N.Y. Acad. Sci. 253, 107ā132.
Rodionov, V. I., Nadezhdina, E. S., Peloquin, J., and Borisy, G. (2001) Digital fluorescence microscopy of cell cytoplasts with and without the centrosome. Methods Cell Biol. 67, 43ā51.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
Ā© 2007 Humana Press Inc.
About this protocol
Cite this protocol
Semenova, I., Rodionov, V. (2007). Fluorescence Microscopy of Microtubules in Cultured Cells. In: Zhou, J. (eds) Microtubule Protocols. Methods in Molecular Medicineā¢, vol 137. Humana Press. https://doi.org/10.1007/978-1-59745-442-1_6
Download citation
DOI: https://doi.org/10.1007/978-1-59745-442-1_6
Publisher Name: Humana Press
Print ISBN: 978-1-58829-642-9
Online ISBN: 978-1-59745-442-1
eBook Packages: Springer Protocols