Near-infrared Fluorescent Probe Research Articles

A Near-Infrared Fluorescent Probe for Visualization of Acetylcholinesterase Flux in the Acute Epileptic Mice Brain

A Near-Infrared Fluorescent Probe for Visualization of Acetylcholinesterase Flux in the Acute Epileptic Mice Brain

Construction of a bifunctional near-infrared fluorescent probe for visualization of copper (II) ions and amyloid-β aggregates in Alzheimer's disease

Alzheimer's disease (AD) is characterized by multiple interconnected pathological factors, including the formation of amyloid-β (Aβ) plaques and the abnormal accumulation of copper (II) ions (Cu2+). The complex interplay between these factors has hindered the development of effective therapeutic agents. The accumulation of Cu2+ not only accelerates Aβ aggregation, but also generates reactive oxygen species through the Fenton reaction, leading to oxidative damage to neurons. While the interaction between Cu2+ and Aβ42 aggregates has been investigated, there is inadequate research into fluorescent probes capable of detecting both Cu2+ and Aβ42 aggregates. This study presents a novel bifunctional near-infrared (NIR) fluorescent probe, LDMD-N, designed using a “Cu2+ activation” strategy. LDMD-N exhibited excellent performance for rapid detection (5 min) and high sensitivity (LOD = 0.0543 µM) of Cu2+ in vitro, and enabled visual fluorescence imaging of fluctuating Cu2+ levels in living cells. Notably, we observed that Cu2+ promoted Aβ42 aggregation in zebrafish using this probe. In vivo and in vitro staining experiments demonstrated that LDMD-N could effectively distinguish AD model mice from wild-type (Wt) mice. This Cu2+ activated bifunctional NIR fluorescent probe provides a promising tool for elucidating the intricate relationship between Aβ42 aggregates and Cu2+. This approach may contribute to a deeper understanding of AD mechanisms and aid in the development of targeted therapeutic strategies.

Novel Colorimetric and Near-Infrared Ratiometric Fluorescent Probe for Sensing Cysteine in Food Samples, Plants, and Living Cells.

Cysteine (Cys) is a crucial biothiol that acts a significant function in food samples and biological systems, including plant roots and living cells. Hence, we developed a novel colorimetric and near-infrared ratiometric fluorescent probe (CT), composed of coumarin and tetrahydroacridine-conjugated indole salt, for the detection of Cys. Upon reaction with Cys, the probe undergoes a specific N-substitution reaction, resulting in a notable colorimetric change and a significant ratiometric fluorescent response in both visible and near-infrared emission channels. These dual-channel ratiometric fluorescence changes are completely independent, enabling the probe to obtain great selectivity, sensitivity, and exceptional detection accuracy. Leveraging these attributes, the probe was employed to provide accurate quantitative analysis of Cys in food samples. Furthermore, confocal imaging demonstrated that the probe could monitor both exogenous and endogenous Cys levels in living cells and track Cys changes in plant roots under heavy metal stress. This work presents a dependable and accurate imaging solution for tracking and identifying Cys of real food, plants, and living cells.

Activatable Near-Infrared Fluorescence Probe for Hypochlorous Acid Detection in Early Diagnosis of Keloids.

Keloids represent pathologic conditions characterized by the presence of hyalinized collagen bundles and chronic inflammatory reactions. Recently, increased ROS production and disrupted apoptosis mechanisms in keloids have been reported, although the detailed mechanisms remain unclear. Herein, we developed a specific fluorescence probe, Pro-NBS, to investigate ClO- levels in keloids. The probe demonstrated high specificity for ClO- over other ROS and exhibited a strong linear detection relationship. Based on its performance, we focused on the TGF-β pathway in the development of keloids. ROS upregulation was observed in keloid-derived fibroblasts. Using ClO- as an intrinsic overexpression marker, our probe effectively distinguished between normal fibroblasts and keloid-derived fibroblasts both in vitro and in vivo. Furthermore, Pro-NBS showed potential for monitoring the progression and evaluating the systematic therapy of abnormal scarring or keloids.

Targeted blood-brain barrier penetration and precise imaging of infiltrative glioblastoma margins using hybrid cell membrane-coated ICG liposomes

Surgical resection remains the primary treatment modality for glioblastoma (GBM); however, the infiltrative nature of GBM margins complicates achieving complete tumor removal. Additionally, the blood-brain barrier (BBB) poses a formidable challenge to effective probe delivery, thereby hindering precise imaging-guided surgery. Here, we introduce hybrid cell membrane-coated indocyanine green (ICG) liposomes (HM-Lipo-ICG) as biomimetic near-infrared (NIR) fluorescent probes for targeted BBB penetration and accurate delineation of infiltrative GBM margins. HM-Lipo-ICG encapsulates clinically approved ICG within its core and utilizes a hybrid cell membrane exterior, enabling specific targeting and enhanced BBB permeation. Quantitative assessments demonstrate that HM-Lipo-ICG achieves BBB penetration efficiency 2.8 times higher than conventional ICG liposomes. Mechanistically, CD44 receptor-mediated endocytosis facilitates BBB translocation of HM-Lipo-ICG. Furthermore, HM-Lipo-ICG enables high-contrast NIR imaging, achieving a signal-to-background ratio of 6.5 in GBM regions of an orthotopic glioma mouse model, thereby improving tumor margin detection accuracy fourfold (84.4% vs. 22.7%) compared to conventional ICG liposomes. Application of HM-Lipo-ICG facilitates fluorescence-guided precision surgery, resulting in complete resection of GBM cells. This study underscores the potential of hybrid cell membrane-coated liposomal probes in precisely visualizing and treating infiltrative GBM margins.Graphical abstract

Imaging nitric oxide dynamics in endoplasmic reticulum stress with a tailor-made near-infrared fluorescence probe

Spatiotemporal fluorescence imaging of NO dynamics during endoplasmic reticulum (ER) stress is crucial for revealing the molecular mechanisms behind NO-mediated neurological and vascular diseases. However, visualizing in vivo NO dynamics during ER stress through fluorescence imaging is challenging due to interference from light scattering and tissue autofluorescence, which limit imaging penetration and reduce the signal-to-noise ratio. Herein, we report the design of a near-infrared fluorescence probe (ER-NO705) for imaging NO dynamics in ER stress. The probe showed an ultrafast and highly selective response manner towards NO, distinguishing it from other biologically relevant species. Notably, ER-NO705 exhibited excellent ER target-ability, which enabled it could be used for spatiotemporal imaging of NO dynamics in ER stress both in living cells, zebrafish and mice. Overall, this study offers a practical imaging agent for studying the role played by NO in ER stress and offering insights into the biological roles of NO in the ER.

Novel BODIPY-based NIR fluorescent probe with appropriate Stokes shift for diagnosis and treatment evaluation of epilepsy via imaging ONOO− fluctuation

Epilepsy is a chronic brain disorder characterized by recurrent seizures that has severely threatened human health. Cumulative research achievements uncover that the pathological progression of epilepsy is closely related to peroxynitrite (ONOO−). In this work, we first constructed BODIPY dye containing pyridine group and indoline group to make it have the donor-acceptor structure, thus achieving NIR emission wavelength (740 nm) and appropriate Stokes shift (80 nm). Based on the BODIPY dye, the novel probe BDP-NIR-Fa was developed that displayed good specificity, faster response speed (60 s), and high signal to noise ratio (247-fold) for monitoring ONOO−. Based on these excellent properties, BDP-NIR-Fa was able to trace endogenous/exogenous ONOO− fluctuations in living cells and mice without interference from other reactive oxygen species. Most importantly, BDP-NIR-Fa could visualize the changes of endogenous ONOO− with remarkable temporal-spatial resolution in KA-induced seizures cell and mice models, that furnishing a potentially forceful tool for diagnosing epilepsy. Finally, BDP-NIR-Fa successfully realized imaging the concentration reduction of ONOO− in epilepsy model during the treatment by an antiepileptic drug. Therefore, the present work provided a reliable approach to investigate the diagnosis and therapy of epilepsy via imaging ONOO− fluctuation.

Renal-Clearable Organic Probes From D-A-D Type Aza-BODIPY Fluorophores for Multiphoton Deep-Brain Imaging.

Bright near-infrared (NIR) fluorescent probes play an important role in in vivo optical imaging. Here, renal-clearable nanodots prepared from Aza-BODIPY are reported fluorophores for multiphoton brain imaging. The design of donor-acceptor-donor (D-A-D) type conjugated structures endowed the fluorophores with large three-photon absorption cross-section for both 1620 and 2200nm excitation. The side chain modification and lipid encapsulation yield ultrasmall nanodots (≈4nm) and a high fluorescence quantum yield (≈0.35) at 720nm emission in the aqueous phase. The measured three-photon action cross-section of a single Aza-BODIPY fluorophore in the nanodots is ≈30 times higher than the commonly used Sulforhodamine 101 dye. Three-photon deep brain imaging of subcortical structures is demonstrated, reaching a depth of 1900µm below the brain surface in a live mouse study. The nanodots enabled blood flow measurement at a depth of 1617µm using line scanning three-photon microscopy (3PM). This work provides superior fluorescent probes for multiphoton deep-brain imaging.

Mitochondria-targeted NIR molecular probe for detecting viscosity of gland damage and SO2 in actual samples

Glandular damage can be caused by various factors, including disease, trauma, or other abnormalities within the organism. The viscosity of the gland is one of the important indicators to measure the degree of damage. Sulfur dioxide (SO2) is widely used as an important food additive due to its preservative and bleaching properties, but its overuse has serious negative impacts on the environment, so it is urgent to develop a simple detection method. Herein, we designed and synthesized a mitochondria-targeted near-infrared (NIR) fluorescence probe (BDC) for the detection of viscosity and SO2. BDC consisted of a donor-π-acceptor (D-π-A) structure and extended double bonds bridging rotor, which enabled sensitive response to viscosity and intense fluorescence emission. The TICT (twisted intramolecular charge transfer) of BDC was inhibited with an increase in viscosity, accompanied by a significant enhancement of red fluorescence signal with emission wavelength beyond 800 nm. Notably, BDC was able to noninvasively and sensitively monitor the viscosity changes in the glands of non-obese diabetic (NOD) mice model. BDC was utilized for monitoring SO2 in food and environmental samples through Michael addition reactions, providing a straightforward tool for SO2 detection in food safety and environmental monitoring.

A near-infrared fluorescent probe with a large Stokes shift for the detection and imaging of biothiols in vitro and in vivo.

In this study, a new near-infrared (NIR) fluorescent turn-on probe featuring a large Stokes shift (198nm) was developed for the detection of biothiols. The probe was based on a dicyanoisophorone derivative serving as the fluorophore and a 2,4-dinitrobenzenesulfonyl (DNBS) group functioning as both a recognition site and a fluorescence quencher. In the absence of biothiols, the fluorescence of the probe was low due to the photoinduced electron transfer (PET) effect between the fluorophore and DNBS. Upon the presence of biothiols, the DNBS group underwent a nucleophilic aromatic substitution reaction with the sulfhydryl group of biothiols, leading to the release of the fluorophore and a notable emission peak at 668nm. This developed probe exhibited exceptional selectivity and sensitivity to biothiols in solution, with an impressive detection limit of 28nM for cysteine (Cys), 22nM for homocysteine (Hcy), and 24nM for glutathione (GSH). Furthermore, the probe demonstrated its applicability by successfully visualizing both endogenous and exogenous biothiols in living systems.

Near-Infrared Ratiometric Hemicyanine Fluorescent Probes for Monitoring Mitochondrial pH Dynamics in Live Cells during Oxidative Stress and Hypoxia.

Novel near-infrared ratiometric molecules (probes A and B) produced by linking formyl-functionalized xanthene and methoxybenzene moieties, respectively, onto a xanthene-hemicyanine framework are detailed. Probe A exhibited a primary absorption peak at 780 nm and a shoulder peak at 730 nm and exhibited fluorescence at 740 nm↓ (signifies a downward shift in intensity upon acidification) in a pH 9.3 buffer and 780 nm↑ at pH 2.8 under excitation at 700 nm. Probe B featured absorptions at 618 and 668 nm at pH 3.2 and at 717 nm at pH 8.6, and fluorescence at 693 nm↑ at pH 3.2 and at 739 nm↓ at pH 8.6, in mostly the red to near-IR region. The ratiometric changes in the intensity of the fluorescent absorptions were reversed between A and B upon acidification as indicated by the arrows. Theoretical calculations confirmed that there were slight changes in conformation between probes and the protonated molecules, suggesting that the changes in emission spectra were due mostly to conjugation effects. Calculations at the APFD/6-311+g(d,p) level with a solvent described by the polarizable continuum model resulted in pK a values for A at 6.33 and B at 6.41, in good agreement with the experimentally determined value of 6.97 and an average of 6.40, respectively. The versatilities of the probes were demonstrated in various experimental contexts, including the effective detection of mitochondrial pH fluctuations. Live cell experiments involving exposure to different pH buffers in the presence of H+ ionophores, monitoring mitophagy processes during cell starvation, studying hypoxia induced by CoCl2 treatment, and investigating responses to various oxidative stresses are detailed. Our findings highlight the potential of attaching xanthene and methoxybenzaldehyde groups onto xanthene-hemicyanine structures as versatile tools for monitoring pH changes in a variety of cellular environments and processes.

A near-infrared fluorescent probe with assembly/aggregation-induced retention effect for specific diagnosis of metastasis and image-guided surgery in breast cancer

Image-guided surgery is crucial for achieving complete tumor resection, reducing postoperative recurrence and improving patient survival. However, current clinical near-infrared fluorescent probes, such as indocyanine green (ICG), face two main limitations: 1) lack of active tumor targeting, and 2) short retention time in tumors, which restricts real-time imaging during surgery. To address these issues, we developed a near-infrared fluorescent probe capable of in situ nanofiber formation within tumor lesions. This probe actively targets the integrin αvβ3 receptors overexpressed on breast cancer cells and exhibits assembly/aggregation-induced retention effects at the tumor site, significantly extending the imaging time window. Additionally, we found that the probe's fluorescence intensity can be enhanced under receptor induction. Due to its excellent tumor specificity and sensitivity, 1FCG-FFGRGD not only identifies primary breast cancer but also precisely locates smaller lymph node metastases and detects sub-millimeter peritoneal metastases. In summary, this near-infrared probe, leveraging assembly/aggregation-induced retention effects, holds substantial potential for various biomedical applications.

Discovery of a novel near-infrared triphenylamine-dicyanoisophorone fluorescent probe with a large Stokes shift for long-term tracking of lysosomes in live cells

Fluorescent probes are powerful tools for real-time tracking of lysosomal movements and spatial distribution, which appears to be essential for understanding the biology of lysosomes. However, the commercially used lysosomal trackers exhibited some limitations, such as narrow Stokes shifts, poor photostability, and green to red instead of near-infrared (NIR) emission. In this study, we have developed a fluorescent probe by incorporating the triphenylamine fragment into the dicyanoisophorone fluorophore. This probe displayed impressive NIR emission (centered at 755 nm) and remarkable Stokes shift (235 nm), which was then demonstrated to be insensitive to lysosomal pH as well as viscosity, and expressed long-term lysosomal tracking ability with little cytotoxicity in live cells. Altogether, this study provided an ideal NIR fluorescent probe for lysosomal tracking.

A high biocompatible near-infrared fluorescent probe for tracking cysteine in multi-biosystem and its application in cervical cancer imaging

Cysteine (Cys) plays a crucial role in the biological system and many related diseases. However, the detection of Cys in living organisms are still hindered by shortage of small molecule fluorophores that exhibit excitation and emission in the near-infrared region. Herein, we designed and synthesized a high water-soluble Cys probe (Cy7-SS) based on heptamethine cyanine scaffold. The prepared Cy7-SS displayed an enhanced emission at near-infrared region (NIR) after the recognition of Cys. Moreover, Cy7-SS not only exhibited high selectivity and sensitivity on the detection of in vitro Cys, but also could be used for endogenous Cys in living cells and C.elegans. The prepared strategy for the design of fluorophores expands the in vivo sensing toolkit for the precise analysis in clinical diagnosis.

The Detection of Lymph Node Metastasis Metastases Using a Near-Infrared Fluorescent Probe Based on Tumor-Targeted Monoclonal Antibody Antibodies Specific: A Prospective Study in a Nude Mouse Model

Background: To identify the metastatic lymph nodes and remove them accurately, the fluorescent surgical navigation ability of the ovarian cancer-specific fluorescent probe COC183B2-800 was assessed to verify the metastatic lymph nodes in the nude mouse model. Methods: The nude mouse model related to lymph node metastases in human ovarian cancer was established using the SKOV3-ip1 cell line. Besides, the COC183B2-800 probe (IRDye800CW Ester conjugated COC183B2 antibody) was fabricated. Moreover, in vivo fluorescence imaging was performed to determine the ability of the COC183B2-800 fluorescent probe to identify metastatic lymph nodes in the nude mouse model. Results: The nude mouse model related to lymph node metastases in human ovarian cancer was successfully established. In vivo fluorescence imaging was performed 30 hours after the injection of the COC183B2-800 fluorescent probe (25 μg) into the animal model, which can achieve specific imaging of metastatic lymph nodes. All metastatic lymph nodes were detected in vivo and in vitro (8/8), and only 1 negative lymph node with reactive enlargement showed a false positive fluorescent signal. Conclusions: The targeted fluorescent probe COC183B2-800 can be employed to identify metastatic lymph nodes in the nude mouse model related to lymph node metastases in human ovarian cancer with high specificity and sensitivity. Targeted fluorescence imaging using COC183B2-800 is expected to become a method to achieve precise lymphadenectomy.

A mitochondria-targeted near-infrared fluorescent probe for pH monitoring in living cells based on the rhodamine-hemicyanine hybrid structure

Mitochondrial pH plays a crucial role in cellular metabolism and pathological conditions. Thus, tracking changes in mitochondrial pH is essential for understanding its impact on cellular processes. In this work, we report a mitochondria-targetable near-infrared pH-sensitive fluorescent probe, Rh-NorCy, based on the rhodamine-hemicyanine hybrid structure. Rh-NorCy contains a non-alkylated indolenine moiety as recognition site of pH and a triphenylphosphonium moiety as the mitochondria-targeting group. As the solution pH decreases from 9.1 to 5.8, the indolium N atom in the Rh-NorCy structure undergoes protonation, leading to a red shift of its maximum absorption wavelength from 568 nm to 709 nm and a significant fluorescence enhancement at 748 nm simultaneously. Importantly, Rh-NorCy exhibits a suitable pKa (7.27) to map the pH variation in the mitochondria. Rh-NorCy demonstrates excellent photostability, minimal cytotoxicity, and strong mitochondria-targeting capability. It has been used to observe mitochondrial pH fluctuations during starvation and carbonylcyanide m-chlorophenylhydrazone (CCCP)-induced mitophagy.

A novel NIR fluorescent probe to image HNO during ferroptosis

BackgroundAs an important reactive nitrogen species (RNS), HNO has been identified as an essential signaling molecule in many physiological processes. Ferroptosis produces a large amount of reactive oxygen species and reactive nitrogen species. However, the detailed mechanism of HNO during process of ferroptosis is rarely reported, especially in the near-infrared range. So, we designed a new near-infrared (NIR) HNO fluorescent probe X-1 based on a tricyanofuran (TCF) derivative and then applied it in ferroptosis imaging. The TCF derivative was chosen as the NIR fluorophore and 2-(diphenylphosphino)benzoate was used as the recognition group. ResultsIn this paper, a novel NIR HNO fluorescent probe X-1 based on tricyanofuran (TCF) derivatives was synthesized using the Staudinger linkage reaction. X-1 exhibited high selectivity for HNO in the near-infrared region (λem = 660 nm). When the recognition group undergoes the Staudinger linkage reaction with HNO, the NIR fluorescence emission increased significantly with the enhancement of the ICT effect. The response mechanism of X-1 to HNO was verified by high-resolution mass spectrometry (HRMS). Probe X-1 has the advantages of fast response (5 min), low detection limit, a large Stokes shift (120 nm) and strong anti-interference ability for HNO recognition. CCK-8 staining result indicates that the probe X-1 has good biocompatibility and little toxic effect on the cells. The probe was successfully applied to imaging the exogenous and endogenous HNO in living cells. SignificanceIn the near-infrared range, HNO was discovered as a mediator of cellular signaling molecules, increasing in concentration during the process of ferroptosis. Furthermore, using this probe, it was further verified that sorafenib, a commonly used drug for cancer treatment, exerts its therapeutic effect by inducing ferroptosis in cancer cells, leading to cell death.

A novel NIR fluorescent probe targeting mitochondria for the detection of Cys using benzothiocarbonate recognition site

Due to the importance of Cys, the development of methods to detect Cys has become a hot research topic. In the present work, we constructed a novel near-infrared fluorescent probe BP-PTC, which specifically binds Cys with a mitochondria-targeting property, using a synthetic benzopyran salt as a chromophore attached to a phenyl thioformate group. The specific response to Cys is achieved based on the addition-cyclisation-elimination reaction process of Cys with phenyl thioformate, allowing it to exhibit fluorescence emission at 670nm. In addition, the fluorescence intensity of BP-PTC shows a good linear relationship with Cys in the concentration range of 0∼28 μΜ. A series of characterization and property tests have demonstrated that BP-PTC has good selectivity and sensitivity for Cys and can be detected by "naked eyes". BP-PTC has been successfully used for the detection of endogenous and exogenous Cys in vivo.

Near-Infrared Fluorescent Turn-On Probe for Selective Detection of Hypochlorite in Aqueous Medium and Live Cell Imaging.

Hypochlorite, as an important reactive oxygen species (ROS), plays a vital role in many physiological and pathological processes, but an excess concentration of hypochlorite (ClO-) may become toxic to humans and cause disease. Hence, the selective and rapid detection of hypochlorite (ClO-) is necessary for human safety. Here, we report a novel near-infrared (NIR) fluorescence "turn-on" and highly selective benzophenoxazinium chloride-based fluorescent probe, BPH (benzophenoxazinium dihydroxy benzaldehyde), for hypochlorite detection. Due to hypochlorite-induced vicinal diol oxidation to the corresponding ortho benzoquinone derivative, the photoinduced electron transfer (PET) process, which was operating from vicinal diol to the benzophenoxazinium chloride receptor moiety, was suddenly inhibited, as a result of which strong NIR fluorescence "turn-on" emission was observed. The detection limit of BPH was found to be 2.39 × 10-10 M, or 0.23 nM. BPH was successfully applied for exogenous and endogenous hypochlorite detection in live MDA-MB 231 cells.

WazaGaY: An Innovative Aza-BODIPY-Derived Near-Infrared Fluorescent Probe for Enhanced Tumor Imaging.

Aza-BODIPYs represent a class of fluorophores in which the π-conjugated system is rigidified and stabilized by a boron atom. A promising strategy to enhance their fluorescence properties involves replacing the boron atom with a metal ion. Here, we describe the synthesis and characterization of a water-soluble derivative where the metal is a gallium(III) ion, termed WazaGaY (water-soluble aza-GaDIPY). Water solubility is ensured by two ammonium substituents, inducing a bathochromic shift and a significant increase in quantum yield compared to that of the dimethylamino analog. The cellular behavior of WazaGaY-1 was observed across different tumor cells. In vivo, the distribution and safety profiles were determined, and tumor uptake was assessed in various tumor types. Following intravenous injection, WazaGaY-1 enabled clear discrimination of tumors engrafted subcutaneously in mice with high tumor-to-muscle ratios (ranging from 7 to 20), even in the absence of specific conjugation. Its potential as a contrast agent for fluorescence-guided surgery was confirmed.