82
C.-L. Hsieh
camera that captures identical COBRI images at a much lower speed (<1000 fps) that
only requires an illumination intensity of <0.01 kW/cm
2 . Under these conditions, no
photo-toxicity is observed.
COBRI microscopy is a simple and powerful imaging modality that provides
high sensitivity and high temporal resolution. COBRI is useful for exploring new cell
dynamics, including the uptake and release of cell vesicles, cell membrane dynamics,
and intracellular or intranuclear dynamics. Research employing COBRI microscopy
requires not only optical observation but also sophisticated data processing and analyses. This makes it an exciting multidisciplinary area combining optics, biophysics,
chemistry, biology, and engineering.
References
1. R.D. Vale, Intracellular transport using microtubule-based motors. Annu. Rev. Cell Biol. 3(1),
347–378 (1987)
2. J. Suh, D. Wirtz, J. Hanes, Efficient active transport of gene nanocarriers to the cell nucleus.
Proc. Natl. Acad. Sci. 100(7), 3878–3882 (2003)
3. N. Hirokawa, S. Niwa, Y. Tanaka, Molecular motors in neurons: transport mechanisms and
roles in brain function, development, and disease. Neuron 68(4), 610–638 (2010)
4. B.W. Guzik, L.S.B. Goldstein, Microtubule-dependent transport in neurons: steps towards an
understanding of regulation, function and dysfunction. Curr. Opin. Cell Biol. 16(4), 443–450
(2004)
5. N. Segev, Coordination of intracellular transport steps by GTPases. Semin. Cell Dev. Biol.
22(1), 33–38 (2011)
6. N. Hirokawa et al., Kinesin superfamily motor proteins and intracellular transport. Nat. Rev.
Mol. Cell Biol. 10, 682 (2009)
7. D. Mudhakir, H. Harashima, Learning from the viral journey: how to enter cells and how to
overcome intracellular barriers to reach the nucleus. AAPS J. 11(1), 65 (2009)
8. R.D. Vale, The molecular motor toolbox for intracellular transport. Cell 112(4), 467–480 (2003)
9. D.J. Stephens, V.J. Allan, Light microscopy techniques for live cell imaging. Science
300(5616), 82–86 (2003)
10. P.L. Leopold et al., Fluorescent virions: dynamic tracking of the pathway of adenoviral gene
transfer vectors in living cells. Hum. Gene Ther. 9(3), 367–378 (1998)
11. I. Wacker et al., Microtubule-dependent transport of secretory vesicles visualized in real time
with a GFP-tagged secretory protein. J. Cell Sci. 110(13), 1453–1463 (1997)
12. M. Lakadamyali et al., Visualizing infection of individual influenza viruses. Proc. Natl. Acad.
Sci. U.S.A. 100(16), 9280–9285 (2003)
13. K. Jaqaman et al., Robust single-particle tracking in live-cell time-lapse sequences. Nat. Methods 5, 695 (2008)
14. N. Ruthardt, D.C. Lamb, C. Bräuchle, Single-particle tracking as a quantitative microscopybased approach to unravel cell entry mechanisms of viruses and pharmaceutical nanoparticles.
Mol. Ther. 19(7), 1199–1211 (2011)
15. S. Manley, J.M. Gillette, J. Lippincott-Schwartz, Chapter 5 Single-particle tracking photoactivated localization microscopy for mapping single-molecule dynamics, in Methods in Enzymology, ed. by N.G. Walter (Academic Press, 2010), pp. 109–120
16. Z. Liu, Luke D. Lavis, E. Betzig, Imaging live-cell dynamics and structure at the singlemolecule level. Mol. Cell 58(4), 644–659 (2015)
17. C. Dietrich et al., Relationship of lipid rafts to transient confinement zones detected by single
particle tracking. Biophys. J. 82, 274–284 (2002)
C.-L. Hsieh
camera that captures identical COBRI images at a much lower speed (<1000 fps) that
only requires an illumination intensity of <0.01 kW/cm
2 . Under these conditions, no
photo-toxicity is observed.
COBRI microscopy is a simple and powerful imaging modality that provides
high sensitivity and high temporal resolution. COBRI is useful for exploring new cell
dynamics, including the uptake and release of cell vesicles, cell membrane dynamics,
and intracellular or intranuclear dynamics. Research employing COBRI microscopy
requires not only optical observation but also sophisticated data processing and analyses. This makes it an exciting multidisciplinary area combining optics, biophysics,
chemistry, biology, and engineering.
References
1. R.D. Vale, Intracellular transport using microtubule-based motors. Annu. Rev. Cell Biol. 3(1),
347–378 (1987)
2. J. Suh, D. Wirtz, J. Hanes, Efficient active transport of gene nanocarriers to the cell nucleus.
Proc. Natl. Acad. Sci. 100(7), 3878–3882 (2003)
3. N. Hirokawa, S. Niwa, Y. Tanaka, Molecular motors in neurons: transport mechanisms and
roles in brain function, development, and disease. Neuron 68(4), 610–638 (2010)
4. B.W. Guzik, L.S.B. Goldstein, Microtubule-dependent transport in neurons: steps towards an
understanding of regulation, function and dysfunction. Curr. Opin. Cell Biol. 16(4), 443–450
(2004)
5. N. Segev, Coordination of intracellular transport steps by GTPases. Semin. Cell Dev. Biol.
22(1), 33–38 (2011)
6. N. Hirokawa et al., Kinesin superfamily motor proteins and intracellular transport. Nat. Rev.
Mol. Cell Biol. 10, 682 (2009)
7. D. Mudhakir, H. Harashima, Learning from the viral journey: how to enter cells and how to
overcome intracellular barriers to reach the nucleus. AAPS J. 11(1), 65 (2009)
8. R.D. Vale, The molecular motor toolbox for intracellular transport. Cell 112(4), 467–480 (2003)
9. D.J. Stephens, V.J. Allan, Light microscopy techniques for live cell imaging. Science
300(5616), 82–86 (2003)
10. P.L. Leopold et al., Fluorescent virions: dynamic tracking of the pathway of adenoviral gene
transfer vectors in living cells. Hum. Gene Ther. 9(3), 367–378 (1998)
11. I. Wacker et al., Microtubule-dependent transport of secretory vesicles visualized in real time
with a GFP-tagged secretory protein. J. Cell Sci. 110(13), 1453–1463 (1997)
12. M. Lakadamyali et al., Visualizing infection of individual influenza viruses. Proc. Natl. Acad.
Sci. U.S.A. 100(16), 9280–9285 (2003)
13. K. Jaqaman et al., Robust single-particle tracking in live-cell time-lapse sequences. Nat. Methods 5, 695 (2008)
14. N. Ruthardt, D.C. Lamb, C. Bräuchle, Single-particle tracking as a quantitative microscopybased approach to unravel cell entry mechanisms of viruses and pharmaceutical nanoparticles.
Mol. Ther. 19(7), 1199–1211 (2011)
15. S. Manley, J.M. Gillette, J. Lippincott-Schwartz, Chapter 5 Single-particle tracking photoactivated localization microscopy for mapping single-molecule dynamics, in Methods in Enzymology, ed. by N.G. Walter (Academic Press, 2010), pp. 109–120
16. Z. Liu, Luke D. Lavis, E. Betzig, Imaging live-cell dynamics and structure at the singlemolecule level. Mol. Cell 58(4), 644–659 (2015)
17. C. Dietrich et al., Relationship of lipid rafts to transient confinement zones detected by single
particle tracking. Biophys. J. 82, 274–284 (2002)