AIE-nanoparticle assisted ultra-deep three-photon microscopy in the<i>in vivo</i>mouse brain under 1300 nm excitation

Li, Dongyu; Zhang, Hequn; Streich, Lina; Wang, Yalun; Lü, Ping; Prevedel, Robert; Qian, Jun

doi:10.1039/d1qm00243k

Cited by 20 publications

(16 citation statements)

References 53 publications

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“…Multiphoton and confocal fluorescence microscopy is advantageous because various fluorescence labeling techniques, together with genetic targeting, allow imaging with high specificity. Previous reports of imaging mouse brains in vivo with two-photon − and three-photon microscopy ,, at depths beyond 1 mm leveraged the optimal wavelength windows for deep tissue penetration at 1300 and 1700 nm. These windows were determined by considering the combined effects of scattering and absorption of ballistic photons in brain tissues in vivo . , On the other hand, one-photon confocal fluorescence microscopy can provide three-dimensionally resolved imaging capabilities and is still widely used in many biology laboratories.…”

mentioning

confidence: 99%

Short-Wave Infrared Confocal Fluorescence Imaging of Deep Mouse Brain with a Superconducting Nanowire Single-Photon Detector

Xia

Gevers²,

Fognini³

et al. 2021

ACS Photonics

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Optical microscopy is a valuable tool for in vivo monitoring of biological structures and functions because of its noninvasiveness. However, imaging deep into biological tissues is challenging due to the scattering and absorption of light. Previous research has shown that the two optimal wavelength windows for high-resolution deep mouse brain imaging are around 1300 and 1700 nm. However, one-photon fluorescence imaging in the wavelength region has been highly challenging due to the poor detection efficiency of currently available detectors. To fully utilize this wavelength advantage, we demonstrated here one-photon confocal fluorescence imaging of deep mouse brains with an excitation wavelength of 1310 nm and an emission wavelength within the 1700 nm window. Fluorescence emission at 1700 nm was detected by a custom-built superconducting nanowire single-photon detector (SNSPD) optimized for detection between 1600 nm and 2000 nm with low detection noise and high detection efficiency. With the PEGylated quantum dots and SNSPD both positioned at the optimal imaging window for deep tissue penetration, we demonstrated in vivo one-photon confocal fluorescence imaging at approximately 1.7 mm below the surface of the mouse brain, through the entire cortical column and into the hippocampus region with a low-cost continuous-wave laser source and low excitation power. We further discussed the significance of the staining inhomogeneity in determining the depth limit of one-photon confocal fluorescence imaging. Our work may motivate the further development of long wavelength fluorescent probes, and inspire innovations in high-efficiency, high-gain, and low-noise long wavelength detectors for biological imaging.

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confidence: 99%

Short-Wave Infrared Confocal Fluorescence Imaging of Deep Mouse Brain with a Superconducting Nanowire Single-Photon Detector

Xia

Gevers²,

Fognini³

et al. 2021

ACS Photonics

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“…2g). The largest σ3 at 1300 nm within the test range was calculated to be 1.26 × 10 -77 cm 6 s 2 , which is the highest value among the reported organic probes 26,29 . Hence, 3PF intensity excited at the 1300-nm wavelength was ~10 times higher than that excited at the 1700-nm wavelength (Fig.…”

Section: The Advantages Of 1300-nm Excitation Window Over 1700-nm Excitation Windowmentioning

confidence: 81%

Ultra-deep through-skull mouse brain imaging via the combination of skull optical clearing and three-photon microscopy

Zhang

et al. 2021

Preprint

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Optical microscopy has enabled in vivo monitoring of brain structures and functions with high spatial resolution. However, the strong optical scattering in turbid brain tissue and skull impedes the observation of microvasculature and neuronal structures at large depth. Herein, we proposed a strategy to overcome the influence induced by the high scattering effect of both skull and brain tissue via the combination of skull optical clearing (SOC) technique and thee-photon fluorescence microscopy (3PM). The Visible-NIR-II compatible Skull Optical Clearing Agents (VNSOCA) we applied reduced the skull scattering and water absorption in long wavelength by refractive index matching and H2O replacement to D2O respectively. 3PM with the excitation in the 1300-nm window reached 1.5 mm cerebrovascular imaging depth in cranial window. Combining the two advanced technologies together, we achieved so far the largest cerebrovascular imaging depth of 1 mm and neuronal imaging depth of >700 μm through intact mouse skull. Dual-channel through-skull imaging of both brain vessels and neurons was also successfully realized, giving an opportunity of non-invasively monitoring the deep brain structures and functions at single-cell level simultaneously.

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“…Owing to the strong fluorescence from DCBT dots and good confinement, the imaging depth of 3PFM imaging can go to 1010 μm, which was among the largest penetration depths of in vivo 3PFM imaging using AIEgens (Figure 4K). 38,48 For better investigating the imaging quality, lines were drawn across tiny capillaries at different imaging depths (e.g., 200, 500, 700, and 1010 μm), and each of the pixel intensities were plotted as a function of position (Figure 4L−O). Gaussian fitting generated from pixel intensity depicted the diameter profile of a capillary, giving rise to a full width at half-maximum (fwhm) of ∼2.8, 3.3, 3.1, and 8.0 μm at depths of 200, 500, 700, and 1010 μm, respectively.…”

Section: Resultsmentioning

confidence: 99%

Intra- and Intermolecular Synergistic Engineering of Aggregation-Induced Emission Luminogens to Boost Three-Photon Absorption for Through-Skull Brain Imaging

et al. 2022

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Three-photon fluorescence microscopic (3PFM) bioimaging is a promising imaging technique for visualizing the brain in its native environment thanks to its advantages of high spatial resolution and large imaging depth. However, developing fluorophores with strong three-photon absorption (3PA) and bright emission that meets the requirements for efficient three-photon fluorescence microscopic (3PFM) bioimaging is still challenging. Herein, four bright fluorophores with aggregation-induced emission features are facilely synthesized, and their powders exhibit high quantum yields of up to 56.4%. The intramolecular engineering of luminogens endows (E)-2-(benzo[d]thiazol-2-yl)-3-(7-(diphenylamino)-9-ethyl-9H-carbazol-2-yl)acrylonitrile (DCBT) molecules with bright near-infrared emission and large 3PA cross sections of up to 1.57 × 10–78 cm6 s2 photon–2 at 1550 nm, which is boosted by 3.6-fold to 5.61 × 10–78 cm6 s2 photon–2 in DCBT dots benefiting from the extensive intermolecular interactions in molecular stacking. DCBT dots are successfully applied for 3PFM imaging of brain vasculature on mice with a removed or intact skull, providing images with high spatial resolution, and even small capillaries can be recognized below the skull. This study will inspire more insights for developing advanced multiphoton absorbing materials for biomedical applications.

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AIE-nanoparticle assisted ultra-deep three-photon microscopy in thein vivomouse brain under 1300 nm excitation

Abstract: Aggregation-induced emission nanoparticles serve as promising fluorescence probes for multi-photon excitation microscopy due to the large absorption cross-section at NIR-IIb region. Here we present organic AIE nanoparticles that feature high...

Cited by 20 publications

References 53 publications

Short-Wave Infrared Confocal Fluorescence Imaging of Deep Mouse Brain with a Superconducting Nanowire Single-Photon Detector

Short-Wave Infrared Confocal Fluorescence Imaging of Deep Mouse Brain with a Superconducting Nanowire Single-Photon Detector

Ultra-deep through-skull mouse brain imaging via the combination of skull optical clearing and three-photon microscopy

Intra- and Intermolecular Synergistic Engineering of Aggregation-Induced Emission Luminogens to Boost Three-Photon Absorption for Through-Skull Brain Imaging

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