Plasmon-enhanced two-photon excitation fluorescence of rhodamine 6G and an Eu-diketonate complex by a picosecond diode laser

Anisotropic noble metal nanoparticles supporting more than one localized surface plasmon resonance can be tailored for efficient dual-mode fluorescence enhancement by ensuring an adequate coupling to both absorption and emission bands of fluorophores. This approach is naturally extended to two-photon excitation fluorescence, where a molecule is excited by simultaneous nonlinear absorption of two photons. However, the relative impact of plasmon coupling to excitation and emission on the overall fluorescence enhancement can be very different in this case. Here, by using the finite-difference time-domain method, we study the two-photon excitation fluorescence of near-infrared fluorescent protein (NirFP) eqFP670, which is the most red-shifted NirFP to date, in proximity to a silver nanobar. By optimizing the length and aspect ratio of the particle, we reach a fluorescence enhancement factor of 103. We show that the single mode coupling regime with highly tuned near-field significantly outperforms the dual-mode coupling enhancement. The plasmon-induced amplification of the fluorophore’s excitation rate becomes of utmost importance due to its quadratic dependence on light intensity, defining the fluorescence enhancement upon two-photon excitation. Our results can be used for the rational design of hybrid nanosystems based on NirFP and plasmonic nanoparticles with greatly improved brightness important for developing whole-body imaging techniques.

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Two-photon patterned photostimulation with low-power, high-efficiency and reliable single-cell optogenetic control

10.1101/2022.01.08.475477 ◽

2022 ◽

Author(s):

Yifan Wang ◽

Yao Zheng ◽

Yongxian Xu ◽

Rongrong Li ◽

Yameng Zheng ◽

...

Keyword(s):

Low Power ◽

Single Cell ◽

Laser Power ◽

High Efficiency ◽

Excitation Density ◽

Two Photon ◽

Photon Excitation ◽

Temporal Focusing ◽

Patterned Illumination ◽

Two Photon Excitation

Two-photon optogenetics enables selectively stimulating individual cells for manipulating neuronal ensembles. As the general photostimulation strategy, the patterned two-photon excitation has enabled millisecond-timescale activation for single or multiple neurons, but its activation efficiency is suffered from high laser power due to low beam-modulation efficiency. Here, we develop a high-efficiency beam-shaping method based on the Gerchberg-Saxton (GS) algorithm with spherical-distribution initial phase (GSSIP) to reduce the patterned two-photon excitation speckles and intensity. It can well control the phase of shaped beams to attain speckle-free accurate patterned illumination with an improvement of 44.21% in the modulation efficiency compared with that of the traditional GS algorithm. A combination of temporal focusing and the GSSIP algorithm (TF-GSSIP) achieves patterned focusing through 500-μm-thickness mouse brain slices, which is 2.5 times deeper than the penetration depth of TF-GS with the same signal-to-noise ratio (SNR). With our method, the laser power can be reduced to only 55.56% of that with traditional method (the temporal focusing with GS, TF-GS) to reliably evoke GCaMP6s response in C1V1-expressing cultured neurons with single-cell resolution. Besides, the photostimulation efficiency is remarkably increased by 80.19% at the same excitation density of 0.27 mW/μm2. This two-photon stimulation method with low-power, reliable and patterned illumination may pave the way for analyzing neural circuits and neural coding and decoding mechanism.

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Stimulated emission in a solution of rhodamine 6G with two-photon excitation

Journal of Applied Spectroscopy ◽

10.1007/bf00609229 ◽

1970 ◽

Vol 12 (6) ◽

pp. 834-834

Author(s):

Yu. A. Bobrovnikov ◽

V. A. Goncharov ◽

G. M. Zverev ◽

A. D. Martynov

Keyword(s):

Rhodamine 6G ◽

Stimulated Emission ◽

Two Photon ◽

Photon Excitation ◽

Two Photon Excitation

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Fluorescence enhancement near metallic nanoparticles under two photon excitation

10.1117/12.500655 ◽

2003 ◽

Cited By ~ 1

Author(s):

Sandrine Leveque-Fort ◽

Emmanuel Fort ◽

Eric Le Moal ◽

Jean Pierre Lacharme ◽

Christian Ricolleau ◽

...

Keyword(s):

Metallic Nanoparticles ◽

Fluorescence Enhancement ◽

Two Photon ◽

Photon Excitation ◽

Two Photon Excitation

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Two-photon excitation of 2,5-diphenyloxazole using a low power green solid state laser

Chemical Physics Letters ◽

10.1016/j.cplett.2010.11.038 ◽

2011 ◽

Vol 501 (4-6) ◽

pp. 572-574 ◽

Cited By ~ 8

Author(s):

Rafal Luchowski

Keyword(s):

Solid State ◽

Low Power ◽

Solid State Laser ◽

State Laser ◽

Two Photon ◽

Photon Excitation ◽

Two Photon Excitation

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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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Two-photon excitation fluorescence microscopy in living systems

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100146618 ◽

1993 ◽

Vol 51 ◽

pp. 154-155 ◽

Cited By ~ 1

Author(s):

David W. Piston

Keyword(s):

Confocal Microscopy ◽

Fluorescence Microscopy ◽

Singlet State ◽

Excited Electronic State ◽

Input Power ◽

Two Photon ◽

Simultaneous Absorption ◽

Photon Excitation ◽

Very High ◽

Two Photon Excitation

Two-photon excitation fluorescence microscopy provides attractive advantages over confocal microscopy for three-dimensionally resolved fluorescence imaging. 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 our fluorescence experiments, the final excited state is the same singlet state that is populated during a conventional fluorescence experiment. Thus, the fluorophore exhibits the same emission properties (e.g. wavelength shifts, environmental sensitivity) used in typical biological microscopy studies. 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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