Bright Near-infrared Emission Research Articles

Bright near-infrared emission from the Au39(SR)29 nanocluster.

The synthesis of atomically precise gold nanoclusters with high photoluminescence quantum yield (PLQY) in the near-infrared (NIR) region and understanding their photoluminescence mechanism are crucial for both fundamental science and practical applications. Herein, we report a highly luminescent, molecularly pure Au39(PET)29 (PET = 2-phenylethanethiolate) nanocluster with PLQY of 19% in the NIR range (915 nm). Steady state and time-resolved PL analyses, as well as temperature-dependent PL measurements reveal the emission nature of Au39(PET)29, which consists of prompt fluorescence (weak), thermally activated delayed fluorescence (TADF), and phosphorescence (predominant). Furthermore, strong dipole-dipole interaction in the solid-state (e.g., Au39(PET)29 nanoclusters embedded in a polystyrene thin-film) is found to narrow the energy gap between the S1 and T1 states, which results in faster intersystem crossing and reverse intersystem crossing; thus, the ratio of TADF to phosphorescence varies and the total PLQY is increased to 32%. This highly luminescent nanocluster holds promise in imaging, sensing and optoelectronic applications.

NIR FRET Luminescence in Rhenium Complex and Dye Co‐Loaded Polymer Nanoparticles

AbstractPhotoactive transition‐metal complexes are luminophores combining high photostability and long luminescence lifetimes. However, reduced optical performance in aqueous solutions has limited their use in biological systems. Herein, the physicochemical and photophysical properties and bioimaging compatibility of Re diimine complexes and near‐infrared (NIR) emitting Cy5 dyes coencapsulated in polymer nanoparticles (NPs) are investigated. By varying the polymers, NPs with sizes from 20 to 70 nm and encapsulating ≤ 40 wt.% of Re complexes, i.e., ≈11 000 Re complexes per NP, are obtained. The photoluminescence (PL) quantum yields of the Re complexes increase eightfold to ≈50% upon encapsulation (vs 6–7% in acetonitrile), resulting in PL brightness up to 108 m−1 cm−1 and PL lifetimes of 3–4 µs. Coencapsulation of Cy5 yields very bright NIR emission upon Re complex excitation. Very close Re‐to‐Cy5 donor–acceptor distances down to ≤2 nm and FRET efficiencies over 90% are confirmed by PL lifetime measurements. The Re‐Cy5 NPs enter mammalian cells for high‐contrast PL imaging in both visible and NIR. This detailed characterization provides a better understanding of the photophysical properties of the transition‐metal‐dye FRET NPs and presents a vital step toward the efficient design of a new class of bright luminescent NP probes.

Versatile side chain turns property and functions of α-cyanostilbene derivatives with near-infrared emission and two-photon property for mitochondrial imaging

Developing red materials with bright near-infrared emission and excellent two-photon performance by a simple preparation process is a meaningful and challenging work. Herein, a series of nitro functionalized D-π-A type α-cyanostilbene derivatives with different side chains (ZH, ZOM, ZOE, and ZOB) have been designed and synthesized by a one-pot method. The compounds show weak emissions and almost the same photophysical properties in solutions. In contrast, in the aggregation state, ZOM, ZOE, and ZOB with side chains of methoxy, ethoxy and benzyloxy emit strong fluorescence, while ZH with no side chain shows weak emission. The aggregate emission mechanism is ascribed to the side chain induced the molecular planarization, and formation of unique “bricks” H-aggregates and “staircase” J-aggregates, investigated by single-crystal analysis and DFT calculations. Considering the excellent photophysical properties of the side chain modified compounds, the triphenylphosphonium is employed to the side chain to obtain a mitochondrial targeting probe Z-TPP, which performs high fluorescence and selectively targeted the mitochondria in live cell and zebrafish under one/two-photon excitation, and further successfully applied to observe the complex mitochondrial network with high signal-to-noise ratio under stimulated emission depletion (STED) nanoscopy.

Bright NIR-Emitting Styryl Pyridinium Dyes with Large Stokes' Shift for Sensing Applications.

Two NIR-emitting donor-π-acceptor (D-π-A) type regioisomeric styryl pyridinium dyes (1a-1b) were synthesized and studied for their photophysical performance and environment sensitivity. The two regioisomers, 1a and 1b, exhibited interesting photophysical properties including, longer wavelength excitation (λex ≈ 530-560 nm), bright near-infrared emission (λem ≈ 690-720 nm), high-fluorescence quantum yields (ϕfl ≈ 0.24-0.72) large Stokes' shift (∆λ ≈ 150-240 nm) and high-environmental sensitivity. Probe's photophysical properties were studied in different environmental conditions such as polarity, viscosity, temperature, and concentration. Probes (1a-1b) exhibited noticeable changes in absorbance, emission and Stokes' shift while responding to the changes in physical environment. Probe 1b exhibited a significant bathochromic shift in optical spectra (∆λ ≈ 20-40 nm) compared to its isomer 1a, due to the regio-effect. Probes (1a-1b) exhibited an excellent ability to visualize bacteria (Bacillus megaterium, Escherichia coli), and yeast (Saccharomyces cerevisiae) via fluorescence microscopy.

ανβ3 integrin-targeted ICG-derived probes for imaging-guided surgery and photothermal therapy of oral cancer.

Indocyanine green (ICG), as the only Federal Drug Administration (FDA) approved fluorescence imaging agent, has been widely applied in clinics for near-infrared (NIR) fluorescence imaging-guided surgery and photothermal therapy of cancers. However, its lack of target specificity and poor photo and photothermal stabilities seriously restrict its wide application in clinical practice. Herein, we developed ICG-derived NIR fluorescent probes consisting of a cypate fluorophore and one or two cyclic-(arginine-glycine-aspartic acid) (cRGD) peptides that can specifically target αvβ3 integrin for accurate diagnosis and therapy of oral tumors. Probe Cy-2RGD has been demonstrated to possess bright NIR emission, great tumor targeting capability and a photothermal effect. Moreover, it could be successfully used for effective imaging-guided surgical resection as well as photothermal therapy of oral tumors. This work could provide a valuable tool for sensitive detection and accurate treatment of malignant tumors.

Near‐Infrared Mechanoluminescence of Cr3+ Doped Gallate Spinel and Magnetoplumbite Smart Materials

AbstractMechanoluminescence (ML), as an optical response to deformation stimuli, shows great potential in high‐end stress sensing, ultrasonic field visualization, and multidimensional anti‐counterfeiting. However, processive practical applications in bio‐medicine are constrained by the discovery of near‐infrared (NIR) ML materials. Unlike lanthanides (Ln3+) with sharp multiplets, two kinds of Cr3+‐doped NIR ML materials, gallate spinel (ZnGa2O4:Cr3+, Zn3Ga2GeO8:Cr3+) and gallate magnetoplumbite (SrGa12O19:Cr3+) are here reported. Owing to the intrinsic cation antisite defects and cation vacancies in the matrix, these materials exhibit bright NIR ML under a relatively low load (20 N). In particular for SrGa12O19:Cr3+ (750 nm, peak; 100 nm, FWHM) with low persistent luminescence (PersL) interference, the ML behavior can be further rejuvenated under UV and sunlight irradiation. SrGa12O19:Cr3+ also shows bright NIR emission under photo‐ and thermo‐stimulation. Owing to their excellent tissue penetration and concealment capability, NIR ML materials show great potential in the fields of bio‐medicine and anti‐counterfeiting.

Tricyano-Methylene-Pyridine Based High-Performance Aggregation-Induced Emission Photosensitizer for Imaging and Photodynamic Therapy.

Photosensitizers equipped with high reactive oxygen species (ROS) generation capability and bright emission are essential for accurate tumor imaging and precise photodynamic therapy (PDT). However, traditional aggregation-caused quenching (ACQ) photosensitizers cannot simultaneously produce desirable ROS and bright fluorescence, resulting in poor image-guided therapy effect. Herein, we report an aggregation-induced emission (AIE) photosensitizer TCM-Ph with a strong donor-π-acceptor (D-π-A) structure, which greatly separates the HOMO-LUMO distribution and reduces the ΔEST, thereby increasing the number of triplet excitons and producing more ROS. The AIE photosensitizer TCM-Ph has bright near-infrared emission, as well as a higher ROS generation capacity than the commercial photosensitizers Bengal Rose (RB) and Chlorine e6 (Ce6), and can effectively eliminate cancer cells under image guidance. Therefore, the AIE photosensitizer TCM-Ph has great potential to replace the commercial photosensitizers.

Access to Pyridinyl or Pyridinium Aza-BODIPYs with Tunable Near-Infrared Fluorescence through ICT from 4-Pyridinyl Pyrroles.

Aza-dipyrromethene boron difluoride (Aza-BODIPY) dyes have attracted much interest in recent decades. In this manuscript, we provide a facile way to access pyridine-substituted pyrroles. A series of 1/7-pyridinyl and pyridinium near-infrared (NIR) Aza-BODIPYs with/without rigidity were constructed and their spectroscopic properties were extensively investigated. These pyridinyl Aza-BOIDIPYs displayed slight bathochromic shifts and maintained bright NIR emission. Pyridinium Aza-BODIPYs showed increased bathochromic shift, however, the molar extinction coefficients and fluorescence quantum yields were decreased owing to ICT effect. The impacts followed the order of pyridin-4-ium>pyridin-2-ium>pyridin-3-ium, which was in consistence with their ICT strength. MO calculation was performed to provide possible explanation to the red-shifted absorption/emission, and apparent charge separation in pyridin-4-ium Aza-BODIPYs. The pyridinium Aza-BODIPY localized in lysosome and potentially avoided harmful ''alkalinizing effects'' of traditional lysosome-targeting fluorescent dyes containing amine moiety. We are working on construction of pyridinyl and pyridinium Aza-BODIPY photosensitizers against microbials or tumors.

A new near-infrared fluorescence indicator derived from chloro-substituted dicyanoisophorone for detecting carboxylesterases (CEs) in living cells and in vivo

Carboxylesterase (CEs) are a kind of liver-synthesized esterase and play a great role in various physiological activities such as drug metabolism and transport processes. Accurate detection of their activity through near-infrared (NIR) fluorescence probe with large Stokes shift is still needed to investigate their biological function and diagnose related disease in vivo. In present study, a new “off-on” fluorescent probe (ZM-1) was constructed with large Stokes shift (160 nm). ZM-1 could be used to detect CEs with a bright NIR emission in a concentration dependent and time-dependent manner. The detection line of ZM-1 is as low as 0.287 × 10−3 U/mL. Moreover, ZM-1 possessed high selectivity and low toxicity. ZM-1 exhibited good optical performance to track CEs activity in cells and monitor the change of CEs activity on 5-Fu-induced model, indicating its good cell membrane permeability and precise detection character. More importantly, ZM-1 was successfully used to in situ image CEs activity in vivo. These studies suggest that probe ZM-1 has potential applicable valuation for CEs detection in living bio-systems and CEs-related disease diagnose.

Electromagnetic Counterparts of Binary-neutron-star Mergers Leading to a Strongly Magnetized Long-lived Remnant Neutron Star

We explore the electromagnetic counterparts that will associate with binary-neutron-star mergers for the case that remnant massive neutron stars survive for ≳0.5 s after the merger. For this study, we employ the outflow profiles obtained by long-term general-relativistic neutrino-radiation magnetohydrodynamics simulations with a mean-field dynamo effect. We show that a synchrotron afterglow with high luminosity can be associated with the merger event if the magnetic fields of the remnant neutron stars are significantly amplified by the dynamo effect. We also perform a radiative transfer calculation for kilonovae and find that, for the highly amplified magnetic field cases, the kilonovae can be bright in the early epoch (t ≤ 0.5 d), while it shows the optical emission which rapidly declines in a few days and the very bright near-infrared emission which lasts for ∼10 days. All these features have not been found in GW170817, indicating that the merger remnant neutron star formed in GW170817 might have collapsed to a black hole within several hundreds milliseconds or magnetic-field amplification might be a minor effect.

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

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.

Water-soluble bright NIR AIEgens with hybrid ROS for wash-free mitochondrial "off-on" imaging and photodynamic therapy.

Several aggregation-induced emission luminogens (AIEgens) with excellent water-solubility and near-infrared emission were designed and synthesized for wash-free "off-on" mitochondrial imaging and photodynamic therapy of HeLa cells. The AIEgen TEPP exhibits both bright near-infrared emission (φF = 17.8%) and high hybrid ROS productivity (including OH˙ and 1O2).

Near-infrared-emitting upconverting BiVO4 nanoprobes for in vivo fluorescent imaging

Near-infrared (NIR) emitting BiVO4:Yb3+,Tm3+ nanoparticles are synthesized by a new solvothermal strategy using solvents of oleic acid and methanol. The obtained BiVO4:Yb3+,Tm3+ samples show an average particle size of ≈164 nm and exhibit an asymmetry monoclinic crystal structure of BiVO4. At NIR excitation of 980 nm, the BiVO4:Yb3+,Tm3+ sample exhibits a nearly single NIR emission at ≈796 nm with extremely weak blue emissions from Tm3+ ions. These high-energy visible emissions are absorbed by the semiconducting host of BiVO4 that possesses a bandgap of ≈2.2 eV. Therefore, the NIR excitation to a single intense NIR emission fluorescent BiVO4 materials could be a potential ideal probe for deep-tissue high-resolution bioimaging. To validate the ability of BiVO4 materials for bio-applications, we conduct the cytotoxicity experiments. The results show that the cytotoxicity of HeLa cells is negligible at a concentration of 0.2 mg/ml of BiVO4:Yb3+,Tm3+ , and the cell viability approaches 90% at a high dosage of 0.5 mg/ml. The Daphnia magna and Zebrafish treated with nanoparticles (0.5 mg/ml) display bright NIR emission without any background, indicating the excellent in vivo fluorescent imaging capacity of BiVO4:Yb3+,Tm3+ nanoparticles. Our findings offer an environment-friendly strategy to synthesize BiVO4 UCL nanophosphors and provide a promising new class of fluorescent probes for biological applications.

High-Performance Near-Infrared Aggregation-Induced Emission Luminogen with Mitophagy Regulating Capability for Multimodal Cancer Theranostics.

The construction of intelligent near-infrared (NIR) fluorophores for high specificity to cancer cells and application in multiple therapeutic modalities is crucial for precise cancer diagnostic and therapy. In this study, an aggregation-induced emission-active NIR fluorophore (TACQ) with mitophagy-modulating activity was synthesized and developed for mitochondrial targeting multimodal cancer theranostics. The strengthened push-pull interaction extended the emission of TACQ into the NIR-II region (>1000 nm). Further, the rotor structure and twisted molecular conformation enables nanoaggregates of TACQ to balance the radiative and nonradiative decays to simultaneously exhibit bright NIR emission, high photothermal conversion efficiency (55%), and efficient generation of reactive oxygen species. The lipocationic property of TACQ allows it to selectively accumulate in the mitochondria of cancer cells. TACQ can induce mitophagy and block mitophagic flux facilitating cancer cell apoptosis. Both in vitro and in vivo evaluations revealed that TACQ is an efficient theranostic agent for NIR fluorescence and photothermal imaging-guided synergistic chemo-photothermal and photodynamic therapy.

Highly Efficient Multifunctional Organic Photosensitizer with Aggregation-Induced Emission for In Vivo Bioimaging and Photodynamic Therapy.

Photosensitizers play a critical role in photodynamic therapy (PDT). Multifunctional organic nanoparticles (NPs) that possess bright fluorescence in aggregates, high singlet oxygen (1O2) quantum yield, near-infrared (NIR) absorption and emission, large Stokes shift, two-photon bioimaging, specific organelle targeting, high PDT efficiency, as well as good biocompatibility and photostability are ideal candidate photosensitizers for image-guided PDT. Due to its enhanced fluorescence and high 1O2 generation efficiency in aggregate states, photosensitizers with aggregation-induced emission (AIE) characteristics have attracted increasing interest in PDT. In this study, a new AIE-active Schiff base 5-(((5-(7-(4-(diphenylamino)phenyl)benzo[c][1,2,5]thiadiazol-4-yl)thiophen-2-yl)methylene)amino)-3-methylthiophene-2,4-dicarbonitrile (TBTDC) based on a D-A-π-A skeleton has been designed and synthesized, and it can be readily encapsulated by Pluronic F-127 to form uniform nanoparticles. TBTDC NPs exhibit bright NIR emission at 825 nm with a Stokes shift up to 300 nm, impressive two-photon bioimaging capability with tissue penetration deep into 300 μm, high 1O2 generation quantum yield (0.552), specific targeting to lysosome, as well as good biocompatibility and photostability. Furthermore, TBTDC NPs present remarkable cytotoxicity for tumor cells and suppression of tumor growth in nude mice through reactive oxygen species generation upon white light irradiation. These results reveal that TBTDC NPs have great potential to become excellent candidates for multifunctional organic photosensitizers for two-photon bioimaging and image-guided PDT and are promising in future clinical applications.

Near-infrared emitting graphene quantum dots synthesized from reduced graphene oxide for in vitro/in vivo/ex vivo bioimaging applications

Near-infrared (NIR) emissive nanomaterials are desired for bioimaging and drug delivery applications due to the high tissue penetration depth of NIR light, enabling in vitro/ex vivo/in vivo fluorescence tracking. Considering the scarcity of NIR-fluorescing biocompatible nanostructures, we have for the first-time synthesized nanometer-sized reduced graphene oxide-derived graphene quantum dots (RGQDs) with NIR (950 nm) emission highly biocompatible in vitro with no preliminary toxic response in vivo. RGQDs are obtained in a high-yield (∼90%) top-down sodium hypochlorite/ultraviolet-driven synthetic process from non-emissive micron-sized reduced graphene oxide (RGO) flakes. This oxidation of RGO yields quantum dots with an average size of 3.54 ± 0.05 nm and a highly crystalline graphitic lattice structure with distinguishable lattice fringes. RGQDs exhibit excitation-independent emission in the visible and NIR-I region with a maximum NIR quantum yield of ∼7%. Unlike their parent material, RGQDs show substantial biocompatibility with ∼75%–80% cell viability up to high (1 mg ml−1) concentrations verified via both MTT and luminescence-based cytotoxicity assays. Tracked in vitro via their NIR fluorescence, RGQDs exhibit efficient internalization in HeLa cells maximized at 12 h with further anticipated excretion. In vivo, RGQDs introduced intravenously to NCr nude mice allow for fluorescence imaging in live sedated animals without the need in sacrificing those at imaging time points. Their distribution in spleen, kidneys, liver, and intestine assessed from NIR fluorescence in live mice, is further confirmed by excised organ analysis and microscopy of organ tissue slices. This outlines the potential of novel RGQDs as NIR imaging probes suitable for tracking therapeutic delivery in live animal models. A combination of smaller size, water-solubility, bright NIR emission, simple/scalable synthesis, and high biocompatibility gives RGQDs a critical advantage over a number of existing nanomaterials-based imaging platforms.

Realizing Near‐Infrared Laser Dyes through a Shift in Excited‐State Absorption (Advanced Optical Materials 6/2021)

The cover image shows organic semiconductor materials for near-infrared lasers, referring to article number 2001947 by Masashi Mamada, Anthony D'Aléo, Chihaya Adachi and co-workers. Based on their core structure, the complete organic emitters are classified as curcuminoids, which are analogues of the curcumin dye found in turmeric. Rational modification of the chemical structure enables bright near-infrared emission and lasing as well as thermally activated delayed fluorescence. The work represents a promising approach for developing materials for near-infrared organic electronic devices.

Highly efficient singlet oxygen generation, two-photon photodynamic therapy and melanoma ablation by rationally designed mitochondria-specific near-infrared AIEgens.

Photosensitizers (PSs) with multiple characteristics, including efficient singlet oxygen (1O2) generation, cancer cell-selective accumulation and subsequent mitochondrial localization as well as near-infrared (NIR) excitation and bright NIR emission, are promising candidates for imaging-guided photodynamic therapy (PDT) but rarely concerned. Herein, a simple rational strategy, namely modulation of donor–acceptor (D–A) strength, for molecular engineering of mitochondria-targeting aggregation-induced emission (AIE) PSs with desirable characteristics including highly improved 1O2 generation efficiency, NIR emission (736 nm), high specificity to mitochondria, good biocompatibility, high brightness and superior photostability is demonstrated. Impressively, upon light irradiation, the optimal NIR AIE PS (DCQu) can generate 1O2 with efficiency much higher than those of commercially available PSs. The excellent two-photon absorption properties of DCQu allow two-photon fluorescence imaging of mitochondria and subsequent two-photon excited PDT. DCQu can selectively differentiate cancer cells from normal cells without the aid of extra targeting ligands. Upon ultralow-power light irradiation at 4.2 mW cm−2, in situ mitochondrial photodynamic activation to specifically damage cancer cells and efficient in vivo melanoma ablation are demonstrated, suggesting superior potency of the AIE PS in imaging-guided PDT with minimal side effects, which is promising for future precision medicine.

Mitochondria-targeting NIR fluorescent probe for rapid, highly sensitive and selective visualization of nitroxyl in live cells, tissues and mice

Nitroxyl (HNO) has been reported to possess unique biological and pharmacological performances, and emerged as a novel therapy for congestive heart failure. Recent studies also suggest that HNO may be produced and involved in important metabolisms in mitochondria. However, due to its high reactivity and short life properties, fast, sensitive and selective observation and monitoring of HNO related dynamic changes in mitochondria still remains a great challenge. Herein, we synthesized a mitochondria-targeting near-infrared (NIR) fluorescent probe (DCMHNO) for rapid detection of HNO with remarkably high sensitivity, selectivity and photostability. DCMHNO shows fast response (about 4 min) towards HNO via 2-(diphenylphosphino)benzoyl group through the Staudinger reaction to boost the bright NIR emission (700 nm) with excellent sensitivity (detection limit of 13 nM), high pH stability and very low interference from other species. DCMHNO can selectively locate in mitochondria and visualize exogenous and endogenous HNO in live HeLa cells with high biocompatibility and photostability. The probe could also monitor the interaction between NO and H2S that gives rise to the generation of HNO in live HeLa cells. In addition, DCMHNO was further utilized in ex vivo NIR imaging of HNO in live mouse liver tissues at the depth of about 50 µm. In vivo imaging of HNO with high signal-to-noise ratio in live mice was also realized by using DCMHNO. These remarkable imaging performances could render NIR DCMNHNO as a useful tool to reveal HNO related dynamic changes in live samples.

Aggregation-Induced Nonlinear Optical Effects of AIEgen Nanocrystals for Ultradeep In Vivo Bioimaging.

Nonlinear optical microscopy has become a powerful tool in bioimaging research due to its unique capabilities of deep optical sectioning, high-spatial-resolution imaging, and 3D reconstruction of biological specimens. Developing organic fluorescent probes with strong nonlinear optical effects, in particular third-harmonic generation (THG), is promising for exploiting nonlinear microscopic imaging for biomedical applications. Herein, a simple method for preparing organic nanocrystals based on an aggregation-induced emission (AIE) luminogen (DCCN) with bright near-infrared emission is successfully demonstrated. Aggregation-induced nonlinear optical effects, including two-photon fluorescence (2PF), three-photon fluorescence (3PF), and THG, of DCCN are observed in nanoparticles, especially for crystalline nanoparticles. The nanocrystals of DCCN are successfully applied for 2PF microscopy at 1040 nm NIR-II excitation and THG microscopy at 1560 nm NIR-II excitation, respectively, to reconstruct the 3D vasculature of the mouse cerebral vasculature. Impressively, the THG microscopy provides much higher spatial resolution and brightness than the 2PF microscopy and can visualize small vessels with diameters of ≈2.7 µm at the deepest depth of 800 µm in a mouse brain. Thus, this is expected to inspire new insights into the development of advanced AIE materials with multiple nonlinearity, in particular THG, for multimodal nonlinear optical microscopy.