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The development of three-photon fluorophores with 1700 nm excitation is pressingly desirable for

Understanding the morphology and hemodynamics of cerebral vasculature at large penetration depths and microscale resolution is fundamentally important to decipher brain diseases. Among the various imaging technologies, three-photon (3P) microscopy is of significance by virtue of its deep-penetrating capability and submicron resolution, which especially benefitsiin vivo/ivascular imaging. Aggregation-induced emission luminogens (AIEgens) have been recognized to be extraordinarily powerful as 3P probes. However, systematic studies on the structure-performance relationship of 3P AIEgens have been seldom reported. Herein, a series of AIEgens has been designed and synthesized. By intentionally introducing benzene rings onto electron donors (D) and acceptors (A), the molecular distortion, conjugation strength, and the D-A relationship can be facilely manipulated. Upon encapsulation with DSPE-PEGsub2000/sub, the optimized AIEgens are successfully applied for 3P microscopy with emission in the far-red/near-infrared-I (NIR-I, 700-950 nm) region under the near-infrared-III (NIR-III, 1600-1870 nm) excitation. Impressively, using mice with an opened skull, vasculature within 1700 μm and a microvessel with a diameter of 2.2 μm in deep mouse brain were clearly visualized. In addition, the hemodynamics of blood vessels were well-characterized. Thus, this work not only proposes a molecular design strategy of 3P AIEgens but also promotes the performance of 3P imaging in cerebral vasculature.

Fluorogens with aggregation-induced emission (AIEgens) are promising optical agents for cellular and vascular imaging. AIEgens with bright emission in the far-red and near-infrared (NIR) region is ...

The photophysical properties of near-infrared-I (NIR-I)-emissive type-II semiconductor nanocrystals (NCs), such as their excitonic properties and multiphoton absorption (MPA), have been rarely repo...

Three-photon excited fluorescence microscopy (3PEFM) has emerged as a promising protocol for visualizing deep-brain vasculature and hemodynamics. However, the current situation is still far from satisfactory, due to small excitation action cross-section and short excitation wavelength of those previously reported 3PEFM luminogens. Herein, we manipulated molecular engineering by subtly regulating structural planarization/twisting to achieve ingenious integration of large three-photon absorption cross-section, high fluorescence quantum yield, ultralong near-infrared IIb excitation, and aggregation-induced emission features. The resulting molecule, namely DPCZ-BT, exhibited as high as 50.6% of fluorescence quantum yield and as large as 2.0 × 10

3-photon microscopy (3PM) excited at the 1700 nm window enables deep-tissue imaging in vivo, especially in brain. PC rod soliton source has previously been exclusively used as the excitation source, which is rather costly and difficult to align. Here we demonstrate a novel nonlinear optical technique to build femtosecond laser source at the 1700 nm window, based on self-phase modulation (SPM) in a short span of large-mode-area fiber. The spectral broadening experienced by the pump pulse leads to the generation of a red-shifted sidelobe at 1603 nm. After spectral filtering, this sidelobe corresponds to 170-fs, 167-nJ pulses at 1603 nm. Using this SPM source, we further demonstrate deep-brain 3 PM to a depth of 1500 μm below the mouse brain surface in vivo. Our SPM femtosecond laser source thus provides a cost effective and easy-to-align alternative excitation source to the PC rod soliton source.

Indocyanine green (ICG) is a human compatible dye and is ideal for deep-tissue two-photon fluorescence (2PF) microscopy excited at the 1700-nm window in vivo. However, the two-photon excitation and emission properties of this dye remain unknown. Here we demonstrate measurement of the two-photon excitation and emission properties of ICG in both water and human plasma, using home-built two-photon action cross-sectional measurement and two-photon emission spectrum measurement systems. Our results show that excited from 1600 to 1800 nm, 2PF can be generated from ICG dissolved in both water and human plasma. The measured two-photon action cross-sectional ησ2 of ICG dissolved in human plasma is an order-of-magnitude larger than that dissolved in water. The measured two-photon emission spectrum overlaps with the one-photon emission spectrum for ICG dissolved in both human plasma and water. Our results will provide key two-photon parameters for the clinical use of ICG.

Myelin is a key component of the peripheral nervous system, whose structure anomaly in the digital skin is implicated in neuropathy. Here we demonstrate an in vivo labeling and imaging technique, capable of visualizing myelin sheaths deep in the mouse digital skin. Through material characterization, we verify that 3-photon fluorescence (3PF) can be generated from a commonly used dye- FluoroMyelin Red for labeling myelin, excited at the 1700-nm window. Topical injection of FluoroMyelin Red in the mouse digit leads to bright labeling of myelin sheaths. Harnessing the deep-penetration capability of 3-photon microscopy excited at the 1700-nm window, we demonstrate that 3PF imaging of FluoroMyelin Red-labeled myelin sheaths in the mouse digit in vivo can be achieved to a depth 340 μm below the skin surface, revealing both branching bundle of and individual myelin sheaths.

Measurement of the hemodynamic physical parameter blood flow speed in the brain in vivo is key to understanding brain physiology and pathology. 2-photon fluorescence microscopy with single blood vessel resolution is typically used, which necessitates injection of toxic fluorescent dyes. Here we demonstrate a label-free nonlinear optical technique, third-harmonic generation microscopy excited at the 1700-nm window, that is promising for such measurement. Using a simple femtosecond laser system based on soliton self-frequency shift, we can measure blood flow speed through the whole cortical grey matter, even down to the white matter layer. Together with 3-photon fluorescence microscopy, we further demonstrate that the blood vessel walls generate strong THG signals, and that plasma and circulating blood cells are mutually exclusive in space. This technique can be readily applied to brain research.

The propagation characteristics of a circular Airy beam (CAB) with Fourier spectrum modified by high-pass, low-pass and band-pass filters (HCAB, LCAB and BCAB for short) are investigated in detail in this paper. Compared with a common CAB, the abruptly autofocusing property of the HCAB is greatly enhanced, while the focal position, focal trajectory and the size of the focal spot do not change. Similar to a CAB with a low-pass filter, a BCAB can form an optical needle but with a much stronger maximum intensity. Not only does the width of this optical needle decrease by half, but also the intensity gradient increases by eight times.

Visualizing deep-brain vasculature and hemodynamics is key to understanding brain physiology and pathology. Among the various adopted imaging modalities, multiphoton microscopy (MPM) is well-known for its deep-brain structural and hemodynamic imaging capability. However, the largest imaging depth in MPM is limited by signal depletion in the deep brain. Here we demonstrate that quantum dots are an enabling material for significantly deeper structural and hemodynamic MPM in mouse brain in vivo. We characterized both three-photon excitation and emission parameters for quantum dots: the measured three-photon cross sections of quantum dots are 4-5 orders of magnitude larger than those of conventional fluorescent dyes excited at the 1700 nm window, while the three-photon emission spectrum measured in the circulating blood in vivo shows a slight red shift and broadening compared with ex vivo measurement. On the basis of these measured results, we further demonstrate both structural and hemodynamic three-photon microscopy in the mouse brain in vivo labeled by quantum dots, at record depths among all MPM modalities at all demonstrated excitation wavelengths.

Elastic fibers are key constituents of the skin. The commonly adopted optical technique for visualizing elastic fibers in the animal skin in vivo is 2-photon microscopy (2 PM) of autofluorescence, which typically suffers from low signal level. Here we demonstrate a new optical methodology to image elastic fibers in animal models in vivo: 3-photon microscopy (3 PM) excited at the 1700-nm window combining with preferential labeling of elastic fibers using sulforhodamine B (SRB). First, we demonstrate that intravenous injection of SRB can circumvent the skin barrier (encountered in topical application) and preferentially label elastic fibers, as verified by simultaneous 2 PM of both autofluorescence and SRB fluorescence from skin structures. Then through 3-photon excitation property characterization, we show that 3-photon fluorescence can be excited from SRB at the 1700-nm window, and 1600-nm excitation is most efficient according to our 3-photon action cross section measurement. Based on these results and using our developed 1600-nm femtosecond laser source, we finally demonstrate 3 PM of SRB-labeled elastic fibers through the whole dermis in the mouse skin in vivo, with only 3.7-mW optical power deposited on the skin surface. We expect our methodology will provide novel optical solution to elastic fiber research.

Fluorescent proteins are widely used to visualize structures and dynamics in various biological samples. Multiphoton microscopy is especially suitable for imaging structures expressing fluorescent proteins with subcellular resolution. 3-photon microscopy (3PM) excited at the 1700-nm window has proven to be promising for deep-tissue (such as brain) imaging expressing red fluorescent proteins. However, the 3-photon excitation and emission spectra of fluorescent proteins suitable at this window remain largely unknown, hampering protein selection and detection optimization. Here we demonstrate detailed measurement of 3-photon excitation and emission spectra for selected fluorescent proteins, suitable for 3-photon excitation at the 1700-nm window. The measured 3-photon excitation spectra will provide guidelines for protein and excitation wavelength selection. The measured 3-photon emission spectra and comparison with the 1-photon emission spectra, on one hand proves that the fundamental Kasha’s rule is still valid for 3-photon fluorescence in these fluorescent proteins, on the other hand will be helpful for efficient fluorescence signal detection.

A boron-dipyrromethene (BODIPY) dye emitting in the near-infrared (NIR) I region (723 nm) exhibits strong saturable absorption at 680 nm. Its multiphoton absorption spectra in the NIR II and III regions are determined. Three-photon fluorescence imaging of the BODIPY-labeled cells excited at 1665 nm is also demonstrated.

The dielectric barrier discharge (DBD) in air at atmospheric pressure is not suitable for industrial applications due to its randomly distributed discharge filaments. In this paper, the influence of the electric field distribution on the uniformity of DBD is theoretically analyzed and experimentally verified. It is found that a certain degree of uneven electric field distributions can control the development of electron avalanches and regulate their transition to streamers in the gap. The discharge phenomena and electrical characteristics prove that an enhanced Townsend discharge can be formed in atmospheric-pressure air with a curved-plate electrode. The spectral analysis further confirms that the gas temperature of the plasma produced by the curved-plate electrode is close to room temperature, which is beneficial for industrial applications. This paper presents the relationship between the electron avalanche transition and the formation of a uniform DBD, which can provide some references for the development and applications of the DBD in the future.

Microglia act as the first and main form of active immune defense in brain. However, in animal models, research on these cells is limited to the superficial layer of the brain, due to the lack of a deep-imaging technique. Here we break this depth limit using three-photon fluorescence (3PF) microscopy excited at the 1700-nm window. Three-photon action cross-section ( η σ 3 ) measurement lays the basis for dye selection and the resultant maximization of 3PF generation. 3PF imaging suppresses the surface background, leading to a much improved signal-to-background ratio compared to the commonly used two-photon microscopy (2PM). We can image microglia 1124 µm below the brain surface in vivo, 3.7 times deeper than previous results using 2PM for microglia imaging. This technique enables us to visualize microglia in the white matter layer in vivo for the first time.

This paper investigates theoretically and numerically the propagation characteristics of a circular Airy beam (CAB) in a uniaxial crystal in detail. The beam loses its boundary cylindrical symmetry during propagation because of the medium anisotropy, although it propagates along the optical axis. This effect of anisotropy on the propagating beam becomes increasingly evident with the increase of the propagation distance. Another main influential factor of the propagation characteristics is the ratio of the extraordinary refractive index to the ordinary refractive index (nsube/sub/nsubo/sub). The more the value deviates from 1, the worse the symmetry of the beam intensity distribution becomes. The polarization becomes notably complicated, but possesses a vortex state with a topological charge of 2 during propagation. The abruptly autofocusing characteristic, the most important property of CABs, also appears when the crystal length is long enough, which is greatly different from that characteristic in isotropic media. This work is helpful for the design of optical devices based on uniaxial crystals for beams with some special wavefronts.

Here we demonstrate deep-brain 2-photon fluorescence microscopy in mouse in vivo, excited at the 1700 nm window. Through signal versus power measurement, we show that indocyanine green (ICG) is a promising 2-photon fluorescent dye excitable at the 1700 nm window. In order to excite ICG circulating in the vasculature in the deep brain, we employ a circular-polarization soliton self-frequency shift technique to generate energetic femtosecond pulses at 1617 nm. Combining the labeling and laser technologies, we achieve a record 2-photon fluorescence brain vasculature imaging depth of 2000 μm in vivo. Both the effective attenuation length measurement and signal-to-background ratio measurement indicate that we have reached the theoretical depth limit in 2-photon fluorescence microscopy.

Here, we demonstrate a polarized high-energy soliton synthesis technique for deep-brain 3-photon microscopy (3PM) excited at the 1700-nm window. Through coherent combining, we generate linearly polarized high-energy solitons whose energy is twice as high than those of each linearly polarized solitons. Due to the nonlinear origin of signals, both measured 3-photon fluorescence signal and third-harmonic signals are thus boosted by ~8 times in a tissue phantom. Using this technique, we further demonstrate 3PM of sulforhodamine 101 labeled vasculature 1600 μm in the mouse brain in vivo, which cannot be achieved by single-polarized soliton excitation.

The propagation characteristics of the circular Airy beam (CAB) modified with low-pass filtering is investigated in details in this paper. Based on a modification of the angular spectrum of CAB, we get a new kind of CAB constructed with low frequency spectrum, which is called “LCAB” for short. A suitable low-pass filter is introduced to cut off the high frequency angular spectrum and maintain the low frequency domain mainly affecting the front light rings of CAB. Two apodization parameters are employed to optimize the low-pass filtering, which influence the propagation characteristic of the LCAB. Fortunately, the abruptly autofocusing property, the most important property of CAB, is still maintained in LCAB. What is more, the initial ring number and the focal spot length can be controlled by adjusting the two apodization parameters. If the two apodization parameters are appropriately chosen, one can form an elegant optical needle which plays an important role in optical manipulations. The numerical results show that the less front light rings are, the longer the optical needle is. It should be noted that the width of the optical needle will increase as long as the length increases, and lead to the decline of the maximum intensity of the optical needle.

We report on the photoresponse of a graphene-metal contact device under terahertz (THz) illumination. The device has an extremely simple structure consisting of a large-area monolayer graphene stripe contacted with two gold electrodes. A significant position-dependent photovoltage is observed across the device by THz excitation, exhibiting a linear relationship with the incident beam power. Experimental results show that the graphene channel length and the substrate thermal conductivity have obvious influence on the photovoltage amplitude and response time, which is consistent with the photothermoelectric mechanism. This compact and powerless device is expected to have a promising application in THz detection. Our work provides theoretical and experimental evidence for the development of high-performance graphene-based THz photodetectors.

We report on the characterization of a terahertz (THz) photodetector with an extremely simple structure consisting of only a macroscopic bundle of double-walled carbon nanotubes (DWCNTs) suspended between two metal electrodes. Polarization-sensitive, broadband, and significant photoresponse occurring at the DWCNT-metal contacts under THz illumination are observed with room-temperature photocurrent and photovoltage responsivities up to ∼16 mA/W and ∼0.2 V/W at 2.52 THz, respectively. Scanning photocurrent measurements provide evidence that the photothermoelectric mechanism dominates the detector response. The simple geometry and compact nature of our device make it suitable for integration and show promising applications for THz detection.