Abstract
We describe a new automatic technique for the study of intracellular mobility. It is based on the visualization of colloidal gold particles by video-enhanced contrast light microscopy (nanometer video microscopy) combined with modern tracking algorithms and image processing hardware. The approach can be used for determining the complete statistics of saltatory motility of a large number of individual moving markers. Complete distributions of jump time, jump velocity, stop time, and orientation can be generated. We also show that this method allows one to study the characteristics of random motion in the cytoplasm of living cells or on cell membranes. The concept is illustrated by two studies. First we present the motility of colloidal gold in an in vitro system of microtubules and a protein extract containing a kinesin-like factor. The algorithm is thoroughly tested by manual tracking of the videotapes. The second study involves the motion of gold particles microinjected in the cytoplasm of PTK-2 cells. Here the results are compared to a study using the spreading of colloidal gold particles after microinjection.
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- Allen R. D., Travis J. L., Allen N. S., Yilmaz H. Video-enhanced contrast polarization (AVEC-POL) microscopy: a new method applied to the detection of birefringence in the motile reticulopodial network of Allogromia laticollaris. Cell Motil. 1981;1(3):275–289. doi: 10.1002/cm.970010302. [DOI] [PubMed] [Google Scholar]
- Allen R. D., Weiss D. G., Hayden J. H., Brown D. T., Fujiwake H., Simpson M. Gliding movement of and bidirectional transport along single native microtubules from squid axoplasm: evidence for an active role of microtubules in cytoplasmic transport. J Cell Biol. 1985 May;100(5):1736–1752. doi: 10.1083/jcb.100.5.1736. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ansorge W. Improved system for capillary microinjection into living cells. Exp Cell Res. 1982 Jul;140(1):31–37. doi: 10.1016/0014-4827(82)90152-5. [DOI] [PubMed] [Google Scholar]
- Axelrod D., Koppel D. E., Schlessinger J., Elson E., Webb W. W. Mobility measurement by analysis of fluorescence photobleaching recovery kinetics. Biophys J. 1976 Sep;16(9):1055–1069. doi: 10.1016/S0006-3495(76)85755-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Barak L. S., Webb W. W. Diffusion of low density lipoprotein-receptor complex on human fibroblasts. J Cell Biol. 1982 Dec;95(3):846–852. doi: 10.1083/jcb.95.3.846. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Brink P. R., Ramanan S. V. A model for the diffusion of fluorescent probes in the septate giant axon of earthworm. Axoplasmic diffusion and junctional membrane permeability. Biophys J. 1985 Aug;48(2):299–309. doi: 10.1016/S0006-3495(85)83783-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
- De Brabander M., Geuens G., Nuydens R., Moeremans M., De Mey J. Probing microtubule-dependent intracellular motility with nanometre particle video ultramicroscopy (nanovid ultramicroscopy). Cytobios. 1985;43(174S):273–283. [PubMed] [Google Scholar]
- De Brabander M., Nuydens R., Geuens G., Moeremans M., De Mey J. The use of submicroscopic gold particles combined with video contrast enhancement as a simple molecular probe for the living cell. Cell Motil Cytoskeleton. 1986;6(2):105–113. doi: 10.1002/cm.970060207. [DOI] [PubMed] [Google Scholar]
- Freed J. J., Lebowitz M. M. The association of a class of saltatory movements with microtubules in cultured cells. J Cell Biol. 1970 May;45(2):334–354. doi: 10.1083/jcb.45.2.334. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gershon N. D., Porter K. R., Trus B. L. The cytoplasmic matrix: its volume and surface area and the diffusion of molecules through it. Proc Natl Acad Sci U S A. 1985 Aug;82(15):5030–5034. doi: 10.1073/pnas.82.15.5030. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hardt S. L. Rates of diffusion controlled reactions in one, two and three dimensions. Biophys Chem. 1979 Nov;10(3-4):239–243. doi: 10.1016/0301-4622(79)85012-7. [DOI] [PubMed] [Google Scholar]
- Jacobson K., Wojcieszyn J. The translational mobility of substances within the cytoplasmic matrix. Proc Natl Acad Sci U S A. 1984 Nov;81(21):6747–6751. doi: 10.1073/pnas.81.21.6747. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kapitza H. G., McGregor G., Jacobson K. A. Direct measurement of lateral transport in membranes by using time-resolved spatial photometry. Proc Natl Acad Sci U S A. 1985 Jun;82(12):4122–4126. doi: 10.1073/pnas.82.12.4122. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Magde D., Elson E. L., Webb W. W. Fluorescence correlation spectroscopy. II. An experimental realization. Biopolymers. 1974 Jan;13(1):29–61. doi: 10.1002/bip.1974.360130103. [DOI] [PubMed] [Google Scholar]
- McCloskey M. A., Poo M. M. Rates of membrane-associated reactions: reduction of dimensionality revisited. J Cell Biol. 1986 Jan;102(1):88–96. doi: 10.1083/jcb.102.1.88. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Murphy D. B., Borisy G. G. Association of high-molecular-weight proteins with microtubules and their role in microtubule assembly in vitro. Proc Natl Acad Sci U S A. 1975 Jul;72(7):2696–2700. doi: 10.1073/pnas.72.7.2696. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Peters R. Nucleo-cytoplasmic flux and intracellular mobility in single hepatocytes measured by fluorescence microphotolysis. EMBO J. 1984 Aug;3(8):1831–1836. doi: 10.1002/j.1460-2075.1984.tb02055.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Salmeen I., Zacmanidis P., Jesion G., Feldkamp L. A. Motion of mitochondria in cultured cells quantified by analysis of digitized images. Biophys J. 1985 Nov;48(5):681–686. doi: 10.1016/S0006-3495(85)83825-X. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Smith B. A., Clark W. R., McConnell H. M. Anisotropic molecular motion on cell surfaces. Proc Natl Acad Sci U S A. 1979 Nov;76(11):5641–5644. doi: 10.1073/pnas.76.11.5641. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Vale R. D., Reese T. S., Sheetz M. P. Identification of a novel force-generating protein, kinesin, involved in microtubule-based motility. Cell. 1985 Aug;42(1):39–50. doi: 10.1016/s0092-8674(85)80099-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wojcieszyn J. W., Schlegel R. A., Wu E. S., Jacobson K. A. Diffusion of injected macromolecules within the cytoplasm of living cells. Proc Natl Acad Sci U S A. 1981 Jul;78(7):4407–4410. doi: 10.1073/pnas.78.7.4407. [DOI] [PMC free article] [PubMed] [Google Scholar]


