Single particle tracking through highly scattering media with multiplexed two-photon excitation

Two-photon excitation has enabled investigators to image living cells in highly scattering media like the central nervous system (1). We have used a custom-built two-photon microscope to image dendritic spines from living cortical pyramidal neurons. Pyramidal cells form the majority of the neuron in the mammalian cortex and they receive practically all their synaptic contacts through dendritic spines. Dendritic spines are small (<1 μm diameter) appendages that have been practically inaccessible to physiological measurements until the application of two-photon excitation to their study (2). We have concentrated in two questions:A- Calcium compartmentalization of spines: Mechanisms of calcium decay kinetics.Dendritic spines can compartmentalize calcium (2). Although the mechanisms of calcium influx into spines have been explored (3), it is unknown what determines the calcium decay kinetics in spines. We investigate calcium dynamics in spines from rat CA1 pyramidal neurons in slices.

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Two-photon single particle tracking in 3D

10.1117/12.274323 ◽

1997 ◽

Cited By ~ 1

Author(s):

Peter T. C. So ◽

Timothy Ragan ◽

Enrico Gratton ◽

Jenny Carerro ◽

Edward Voss

Keyword(s):

Particle Tracking ◽

Single Particle ◽

Single Particle Tracking ◽

Two Photon

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NIR-emitting molecular-based nanoparticles as new two-photon absorbing nanotools for single particle tracking

10.1117/12.2189638 ◽

2015 ◽

Cited By ~ 1

Author(s):

J. Daniel ◽

A. G. Godin ◽

G. Clermont ◽

B. Lounis ◽

L. Cognet ◽

...

Keyword(s):

Particle Tracking ◽

Single Particle ◽

Single Particle Tracking ◽

Two Photon

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Two-photon excitation microscopy in cellular biophysics

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100136684 ◽

1995 ◽

Vol 53 ◽

pp. 62-63

Author(s):

David W. Piston ◽

Brian D. Bennett ◽

Robert G. Summers

Keyword(s):

Confocal Microscopy ◽

Laser Beam ◽

Duty Cycle ◽

Excited Electronic State ◽

Input Power ◽

Two Photon ◽

Simultaneous Absorption ◽

Photon Excitation ◽

Very High ◽

Two Photon Excitation

Two-photon excitation microscopy (TPEM) provides attractive advantages over confocal microscopy for three-dimensionally resolved fluorescence imaging and photochemistry. Two-photon excitation arises from the simultaneous absorption of two photons in a single quantitized event whose probability is proportional to the square of the instantaneous intensity. For example, two red photons can cause the transition to an excited electronic state normally reached by absorption in the ultraviolet. In practice, two-photon excitation is made possible by the very high local instantaneous intensity provided by a combination of diffraction-limited focusing of a single laser beam in the microscope and the temporal concentration of 100 femtosecond pulses generated by a mode-locked laser. Resultant peak excitation intensities are 106 times greater than the CW intensities used in confocal microscopy, but the pulse duty cycle of 10-5 maintains the average input power on the order of 10 mW, only slightly greater than the power normally used in confocal microscopy.

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