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With suppressed photon scattering and diminished autofluorescence, in vivo fluorescence imaging in the 1,500- to 1,700-nm range of the near-IR (NIR) spectrum (NIR-IIb window) can afford high clarity and deep tissue penetration. However, there has been a lack of NIR-IIb fluorescent probes with sufficient brightness and aqueous stability. Here, we present a bright fluorescent probe emitting at ∼1,600 nm based on core/shell lead sulfide/cadmium sulfide (CdS) quantum dots (CSQDs) synthesized in organic phase. The CdS shell plays a critical role of protecting the lead sulfide (PbS) core from oxidation and retaining its bright fluorescence through the process of amphiphilic polymer coating and transferring to water needed for imparting aqueous stability and compatibility. The resulting CSQDs with a branched PEG outer layer exhibited a long blood circulation half-life of 7 hours and enabled through-skin, real-time imaging of blood flows in mouse vasculatures at an unprecedented 60 frames per second (fps) speed by detecting ∼1,600-nm fluorescence under 808-nm excitation. It also allowed through-skin in vivo confocal 3D imaging of tumor vasculatures in mice with an imaging depth of ∼1.2 mm. The PEG-CSQDs accumulated in tumor effectively through the enhanced permeation and retention effect, affording a high tumor-to-normal tissue ratio up to ∼32 owing to the bright ∼1,600-nm emission and nearly zero autofluorescence background resulting from a large ∼800-nm Stoke’s shift. The aqueous-compatible CSQDs are excreted through the biliary pathway without causing obvious toxicity effects, suggesting a useful class of ∼1,600-nm emitting probes for biomedical research.

Real-time imaging of the tumour boundary is important during surgery to ensure that sufficient tumour tissue has been removed. However, the current fluorescence probes for bioimaging suffer from poor tumour specificity and narrow application of the imaging window used. Here, we report a bioactivated in vivo assembly (BIVA) nanotechnology, demonstrating a general optical probe with enhanced tumour accumulation and prolonged imaging window. The BIVA probe exhibits active targeting and assembly induced retention effect, which improves selectivity to tumours. The surface specific nanofiber assembly on the tumour surface increases the accumulation of probe at the boundary of the tumor. The blood circulation time of the BIVA probe is prolonged by 110 min compared to idocyanine green. The assembly induced metabolic stability broaden the difference between the tumor and background, obtaining a delayed imaging window between 8–96 h with better signal-to-background contrast (>9 folds). The fabricated BIVA probe permits precise imaging of small sized (<2 mm) orthotopic pancreatic tumors in vivo. The high specificity and sensitivity of the BIVA probe may further benefit the intraoperative imaging in a clinical setting. Fluorescence probes for detecting tumours during surgery can suffer from poor accumulation and short imaging windows. Here, the author develop fluorescence probes with multiple motifs that permit enhanced circulation times, tumour targeting and use the probes to image pancreatic cancer in mice

Carbon dots (CDs) are a strong alternative to conventional fluorescent probes for cell imaging due to their brightness, photostability, tunable fluorescence emission, low toxicity, inexpensive preparation, and chemical diversity. Improving the targeting efficiency by modulation of the surface functional groups and understanding the mechanisms of targeted imaging are the most challenging issues in cell imaging by CDs. Firstly, we briefly discuss important features of fluorescent CDs for live-cell imaging application in this review. Then, the newest modulated CDs for targeted live-cell imaging of whole-cell, cell organelles, pH, ions, small molecules, and proteins are elaborately discussed, and their challenges in these fields are explained. Graphical abstract

N-rich metal-free and metal-doped carbon quantum dots (CQDs) have been prepared through one-step hydrothermal method using tetraphenylporphyrin or its transition metal (Pd or Pt) complex as precursor. The structures and morphology of the as-prepared nanoparticles were analyzed by X-ray diffraction, high-resolution transmission electron microscopy, X-ray photoelectron spectroscopy, and Fourier transform infrared spectra. Three kinds of nanocomposites show similar structures except for the presence of metal ions in Pd-CQDs and Pt-CQDs indicated by X-ray photoelectron spectroscopy. All of them display bright blue emission upon exposure to ultraviolet irradiation. The CQDs exhibit typical excitation-dependent emission behavior, with the emission quantum yield of 10.1%, 17.8%, and 15.2% for CQDs, Pd-CQDs, and Pt-CQDs, respectively. Moreover, the CQDs, Pd-CQDs, and Pt-CQDs could serve as fluorescent probes for the specific and sensitive detection of Fe ions in aqueous solution. The low cytotoxicity of CQDs is demonstrated by MTT assay against HeLa cells. Therefore, the CQDs can be used as efficient probes for cellular multicolor imaging and fluorescence sensors for the detection of Fe ions due to their low toxicity, excellent biocompatibility, and low detection limits. This work provides a new route to synthesize highly luminescent N-rich metal-free or metal-doped CQDs for multifunctional applications.

A rapid, sensitive, and selective fluorometric assay is described for the determination of chromium(VI) in real waters and living cells. The method is making use of nitrogen, phosphorus, and sulfur tri-doped carbon dots (NPS-CDs) which have absorption/emission maxima at 360/505 nm/nm. Cr(VI) has an absorption maximum at 350 nm and causes an inner filter effect (IFE) on the blue fluorescence of the NPS-CDs. The NPS-CDs were hydrothermally synthesized using p -aminobenzenesulfonic acid and tetrakis(hydroxymethyl)phosphonium chloride as precursors. The NPS-CDs were characterized by transmission electron microscopy, X-ray diffraction, and several spectroscopic methods. They are biocompatible and negligibly cytotoxic when tested with HeLa cells and MCF-7 cells even after 48 h of incubation. The NPS-CDs were used as fluorescent probes for Cr(VI). The detection limit is 0.23 μM (three times standard deviation versus slope), and the linear response covers the 1 to 500 μM chromate concentration range. The NPS-CDs were applied to the determination of Cr(VI) in real waters and living cells (HeLa and MCF-7) and gave satisfying results. Graphical abstract Schematic representation of hydrothermal synthesis of nitrogen, phosphorus, and sulfur tri-doped carbon dots (NPS-CDs) for Cr(VI) detection via inner filter effect (IFE). NPS-CDs were applied to the determination of Cr(VI) in living cells (HeLa and MCF-7) with satisfying results.

Aminoglycosides (AGs) are broad-spectrum antibiotics that are associated with kidney damage, balance disorders, and permanent hearing loss. This damage occurs primarily by killing of proximal tubule kidney cells and mechanosensory hair cells, though the mechanisms underlying cell death are not clear. Imaging molecules of interest in living cells can elucidate how molecules enter cells, traverse intracellular compartments, and interact with sites of activity. Here, we have imaged fluorescently labeled AGs in live zebrafish mechanosensory hair cells. We determined that AGs enter hair cells via both nonendocytic and endocytic pathways. Both routes deliver AGs from the extracellular space to lysosomes, and structural differences between AGs alter the efficiency of this delivery. AGs with slower delivery to lysosomes were immediately toxic to hair cells, and impeding lysosome delivery increased AG-induced death. Therefore, pro-death cascades induced at early time points of AG exposure do not appear to derive from the lysosome. Our findings help clarify how AGs induce hair cell death and reveal properties that predict toxicity. Establishing signatures for AG toxicity may enable more efficient evaluation of AG treatment paradigms and structural modifications to reduce hair cell damage. Further, this work demonstrates how following fluorescently labeled drugs at high resolution in living cells can reveal important details about how drugs of interest behave.

Red emissive B,N co-doped carbon dots (BN-CDs) were hydrothermally synthesized from cresyl violet and boric acid. The BN-CDs exhibited excellent photostability, low cytotoxicity, excitation/emission maxima at 520/616 nm, and a relatively high quantum yield of 18%. The BN-CDs can binded to mercury(II), and this results in quenching of the red-colored fluorescence. However, on subsequent addition of the biothiol (such as cysteine, homocysteine or glutathione), fluorescence recovers. Therefore, the BN-CDs can be used as a multifunctional probe based on “on-off-on” fluorescence response for the detection of Hg(II) and biothiols. The following detection limits were accomplished: (a) Hg(II): 2.8 μM; (b) glutathione: 1.7 μM; (c) cysteine: 2.3 μM; (d) homocysteine: 3.0 μM. The BN-CDs also have been successfully applied for the imaging of Hg(II) and biothiols in HepG2 cells with excellent bio-compatibility. Graphical abstract Red emissive B,N co-doped carbon dots (BN-CDs) were synthesized through hydrothermal treatment of cresyl violet and boric acid. The BN-CDs can be used as a multifunctional probe based on “on-off-on” fluorescence response for detecting mercury(II) and biothiols in aqueous solution and living cells.

The development of fluorophores with high-fluorescence quantum yields is highly desirable. To regulate photophysical properties, previous fumaronitrile-core fluorophore designs primarily employed electron-donating structure modifications and π-conjugation extension strategies. Here, we report a novel strategy to enhance the fluorescence performance of fluorophores by introducing methyl groups into fumaronitrile phenyl rings. The introduction of methyl groups reduces the ability to generate reactive oxygen species while enhancing the fluorescence quantum yield. Notably, after encapsulating DSPE-PEG2000 to form nanoparticles, TFN-Me nanoparticles exhibited superior fluorescence performance than previously reported fluorophores and successfully applied in in vivo tumor fluorescence imaging. This study indicates that the methyl introduction strategy holds the potential to become a powerful tool for developing high-brightness fluorophores with fumaronitrile structure.

Tunable multicolor carbon dots (CDs) with a quantum yield reach up to 35% were generated directly from rhodamine and urea via one-step hydrothermal approach and purified through silica gel column chromatography. Transmission electron microscopy images reveal that the as-prepared CDs possess a small size distribution below 10 nm with bright blue, green, and yellow color emission, designated as b-CDs, g-CDs, and y-CDs, respectively. The in-depth investigations reveal that the multicolor emission CDs with different fraction displays fluorescence emission wavelength ranges from 398 nm (b-CDs), 525 nm (g-CDs), to 553 nm (y-CDs) which could be well modulated by controlling the amount of heteroatom nitrogen especially amino nitrogen onto their surface structures. Further experiments verify the important role of nitrogen content by using rhodamine solely or substituting urea with sulfur containing compounds as precursors to produce corresponding CDs since the performance is lower than that of urea incorporation. Theoretical calculation results also reveal that the increasing amount of amino nitrogen into their surface structures of b-CDs, g-CDs to y-CDs is responsible for reduced band gaps energy, which result in the redshifted wavelength. Benefiting from the excellent photoluminescence properties, wide pH variation range, high photo stability, and low toxicity, these CDs were employed for HClO sensing at 553 nm within the range 5 to 140 μM with a limit of detection (LOD) of 0.27 ± 0.025 μM ( n  = 3) and multicolor cellular imaging in HeLa cells. Graphical abstract Tunable multicolor carbon dots (CDs) were generated directly from rhodamine and urea via one-step hydrothermal approach and purified through silica gel column chromatography. The as-prepared CDs exhibit bright blue, green, and yellow color emission which could be well modulated by controlling the increasing incorporation of heteroatom nitrogen especially amino nitrogen into their surface structures. These CDs were employed for HClO sensing and demonstrated to multicolor cellular imaging in HeLa cells.

New green-emissive carbon dots (G-CDs) are described here and shown to be viable fluorescent nanoprobes for the detection of changes in cellular pH values. By using m -phenylenediamine as the carbon source, G-CDs with an absolute quantum yield of 36% were solvothermally synthesized in the presence of strong H 2 SO 4 . The G-CDs have an average size of 2.3 nm and display strong fluorescence with excitation/emission peaks at 450/510 nm. The fluorescence intensity depends on the pH value in the range from 6.0 to 10.0, affording the capability for sensitive detection of intracellular pH variation. The nanosensor with excellent photostability exhibited good fluorescence reversibility in different pH solutions, and showed excellent stability against the influence of other biological species. The nanoprobe was successfully used in confocal fluorescence microscopy to determine pH values in SMMC-7721 cells. Graphical abstract Schematic presentation of green-emissive carbon dots (G-CDs) synthesized using m -phenylenediamine and sufuric acid through a solvothermal method for real-time fluorometric monitoring of intracellular pH values. Mechanism can be ascribed to PET process from the electron lone pair in amino group to the CDs.