Dual-response Fluorescent Probe Research Articles

A dual-response fluorescent hemicyanine probe for the detection of mitochondrial hypochlorite and viscosity based on ESIPT/AIE and TICT

Viscosity and hypochlorite (ClO−) are two important factors affecting the physiological activity and function of mitochondria. Hypochlorite (ClO−) is a vital reactive oxygen species, and endogenous ClO− acts as an important germicidal oxidant in the immune system at low concentrations. While, fluctuations in viscosity can cause mitochondrial dysfunction that is linked to several diseases. We herein report on a hemicyanine-based probe (BDTB-PA) for the detection of hypochlorite (ClO−) and viscosity. The probe exhibits a very low fluorescence background in an aqueous solution due to twisted intramolecular charge transfer (TICT) and blocked excited-state intramolecular proton transfer (ESIPT). In a viscous solution, the TICT of the probe was inhibited, generating a strong emission at 580 nm. In contrast, upon treatment with ClO−, the phenylboronic acid pinacol ester group of probe BDTB-PA was removed, and the hydroxyl group was released, activating the ESIPT process. Coupled with its aggregation-induced emission (AIE) characteristics, the product of BDTB-PA with ClO− exhibited a distinct emission at 540 nm. As such a solution of the probe BDTB-PA exhibited a distinct fluorescent color change from colourless to green under UV light with the addition of ClO−. More importantly, probe BDTB-PA was used for monitoring ClO− and viscosity in the mitochondria of living cells, plants and zebrafish.

A dual-responsive fluorescent probe for cellular hypoxia and tumor imaging based on nitroreductase and viscosity synergy

Nitroreductase (NTR), an enzyme that operates within the cytoplasm, relies on flavin mononucleotide or flavin adenine dinucleotide for its activity. In tumor cells, hypoxic conditions often lead to an increase in NTR levels. Consequently, the development of fluorescent probes capable of detecting NTR activity is crucial for the monitoring of hypoxia in biological systems. This study presents the one-step synthesis of a near-infrared fluorescent probe, DSM-NO2. The probe was highly sensitive to NTR and caused a fluorescence enhancement by 50-fold, with a detection limit as low as 1 ng/mL. Under hypoxic conditions in tumor cells, not only does NTR increase, but the cellular viscosity also rises. In high-viscosity environments, the fluorescence enhancement could reach up to 220-fold, and the detection limit could be further reduced to 55 pg/mL. The reaction mechanism was thoroughly investigated using mass spectrometry, molecular docking, and theoretical calculations. Moreover, the probe was successfully applied in the real-time monitoring and visualization analysis of NTR during the growth of Escherichia coli and in the hypoxic state of cancer cells. Additionally, through nebulized inhalation, the probe was successfully employed for in situ fluorescent imaging studies in a murine model of lung cancer.

Dual-response fluorescent probe for lipid droplets and hypochlorous acid and imaging study in inflammation and atheromatous plaques

Simultaneously detecting two biomarkers can significantly enhance disease diagnosis accuracy and deepen our understanding of disease pathogenesis. Developing a single fluorescent probe with two independent response units for this purpose is crucial. In line with this, a novel fluorescent probe (TPA-DT-DNH) had been proposed, featuring triphenylamine and C=N groups sensitive to lipid droplets (LDs) and hypochlorous acid (HClO), respectively. Spectral results highlighted that only the co-presence of LDs and HClO could “turn-on” the fluorescence of TPA-DT-DNH, when the concentration of one substance remained constant, the fluorescence intensity linearly increased with the concentration of the other substance. The imaging results demonstrated that TPA-DT-DNH can differentiate between normal cells and cells rich in LDs when HClO was present, and can detect HClO in HeLa cells. Furthermore, dynamic changes in the inflammation process induced by LPS can be visualized in both cells and zebrafish. Additionally, significant fluorescence enhancement was observed in atheromatous plaque.

A single-molecule fluorescent probe for visualizing viscosity and hypoxia in lysosomes and zebrafish embryos.

Viscosity and hypoxia, as microenvironment parameters, play important roles in maintaining normal biological processes and homeostasis. Therefore, simultaneous and sensitive detection of these elements with simple and effective methods could offer precise information in biology. Here, we report a two-site lysosome-targeting fluorescent probe, NVP, for monitoring viscosity and nitroreductase with dual emission channels (emission shift is 86 nm). The NVP probe has displayed highly sensitive and selective responses towards viscosity and nitroreductase, respectively. Significantly, the fluctuations of viscosity and NTR have been detected in vitro and in vivo. We expect that the dual-responsive fluorescent NVP probe will become a potential molecular tool for the exploration of deeper functions of viscosity and hypoxia.

A mitochondria-targeted dual-response fluorescent probe with large Stokes shift for visualizing viscosity and cysteine in cancer cells and zebrafish

Mitochondrial cysteine (Cys) and viscosity are closely associated with numerous cellular processes and diseases. Developing a method capable of real-time tracking of viscosity and Cys in mitochondria is crucial for understanding cell status and mitochondria-related diseases. In this study, we developed a mitochondria-targeted fluorescent probe (Mito-QP) with a large Stokes shift of 171 nm for detecting Cys and viscosity. The acryloyl group enabled Mito-QP specific response to Cys with high sensitivity. The probe exhibited significant red fluorescence enhancement with increasing viscosity due to restricted intramolecular rotation. Additionally, Mito-QP showed low cytotoxicity and excellent mitochondrial-targeting ability. The probe was successfully employed to monitor fluctuations in mitochondrial viscosity and Cys in cancer cells and zebrafish. This work provides a convenient and effective sensor for studying mitochondrial dysfunction.

Design, synthesis and application of polarity- and pH-sensitive dual-responsive small molecule fluorescent probe for early cancer diagnosis

Cancer is one of the deadliest and most complex diseases globally. It is well known that cancer cells exhibit a unique intracellular microenvironment characterized by low pH and low polarity compared to normal cells. Thus, dual-responsive fluorescent probe are more suitable for such complex and variable physiological environments than single-response probe. In this study, we designed and synthesized a dual-responsive fluorescent probe, CCT, which responds to both pH and polarity based on intramolecular charge transfer (ICT) and photoelectron transfer (PET) mechanisms.We synthesized probe CCT by combining naphthylimide, a coumarin unit, and morpholine through a six-step process. The PET mechanism between morpholine and naphthylimide renders probe CCT pH-sensitive. Additionally, the direct linkage of the polarity-sensitive units, coumarin and naphthylimide, creates a larger polarity-sensitive system. When both low pH and low polarity conditions are present, the fluorescence intensity of probe CCT is significantly higher than under separate conditions. This increased fluorescence intensity matches the low pH and low polarity microenvironment of cancer cells, facilitating bioimaging.Fluorescence imaging results demonstrate that probe CCT effectively distinguishes between cancer and normal cells, suggesting its potential as a powerful tool for early cancer diagnosis.

A dual-response fluorescent probe for detecting sulfur dioxide and hypochlorous acid in cells, water and food

Hypochlorous acid (HClO) and sulfur dioxide (SO2) play vital roles in various fields, including living organisms, water samples, food and so on. High doses of SO2 and HClO are seriously harmful to human and are associated with a variety of diseases. Therefore, monitoring the level of SO2 and HClO in various fields is significant for exploring their roles. In this work, a novel dual-response fluorescent probe RPT for detecting SO2 and HClO is successfully designed and synthesized, which is composed by pyrylium moiety and thiazine part. The fluorescent probe RPT can respectively monitor SO2 and HClO by two well-separated emission wavelength with an obvious difference of 85 nm. The fluorescent probe RPT also displays desired properties such as simple synthetic method, high sensitivity and selectivity. In addition, as a biocompatible molecule, RPT has been successfully used for monitoring SO2 and HClO in HeLa cells and RAW 264.7 cells in two different channels (red channel and green channel). More importantly, SO2 and HClO levels in various water and food samples have been detected with satisfactory results.

A dual-response fluorescent probe to reveal the role of ferroptosis in drug-induced liver injury

Drug-induced liver injury (DILI) has already been recognized as a formidable obstacle in global health issues. Despite the proposed common cellular death types in DILI, exploration of alternative mechanisms such as ferroptosis for prevention and therapy remains limited. Visualization of the role of ferroptosis in DILI holds profound promise in advancing diagnostic and therapeutic strategies for DILI. To address this, herein, a unique multi-functional fluorescent probe, VIS-HNO, was elaborately built up and endowed with the extraordinary performance of excellent selectivity and high sensitivity toward HNO and viscosity. Employing the probe VIS-HNO with the confocal fluorescence imaging, the up-regulations of HNO and viscosity in the process of DILI and ferroptosis were disclosed. Then the remediation of DILI was achieved by VIS-HNO under the treatment of ferrostatin1 (Fer-1), indicating its therapeutic value for DILI. In addition, taking into account the significance of HNO in determining the role of ferroptosis in DILI, the pathway of HNO production in DILI was further explored, which presented essential insights into the role of ferroptosis in DILI. Overall, these findings demonstrated that VIS-HNO may be a potent tool to aid in diagnosing ferroptosis and DILI, more importantly, to understand the role of ferroptosis in DILI by evaluating HNO and viscosity level, which will facilitate the exploration of unique strategies for DILI treatment.

An ESIPT + AIE based dual-response fluorescent probe for continuous detection of PhSH and HClO and visualization of PhSH-induced oxidative stress in living cells

As a valuable industrial chemical, thiophenol (PhSH) is poisonous, which can be easily absorbed by the human body, leading to many serious health issues. In addition, PhSH-triggered oxidative stress is considered to be related with the pathogenesis and toxicity of PhSH. Therefore, efficient methods for monitoring PhSH and ROS production induced by PhSH in living systems are very meaningful and desired. Herein, we reasonably developed a facile dual-response fluorescent probe (HDB-DNP) by incorporating the dinitrophenyl (DNP) group into a novel methylthio-substituted salicylaldehyde azine (HDB) with AIE and ESIPT features. The probe itself was non-fluorescent owing to the strong quenching effect of DNP group. In the presence of PhSH, HDB-DNP gave an intense red fluorescence (610 nm), which can rapidly switch to green fluorescence (510 nm) upon further addition of HClO, allowing the successive detection of PhSH and HClO in two well-separated channels. HDB-DNP proved to be a very promising dual-functional probe for rapid (PhSH: < 17 min; HClO: 10 s) and selective detection of PhSH and HClO in physiological conditions with low detection limit (PhSH: 13.8 nM; HClO: 88.6 nM). Inspired by its excellent recognition properties and low cytotoxicity, HDB-DNP was successfully applied for monitoring PhSH and PhSH-induced HClO generation in living cells with satisfactory results, which may help to better understand the pathogenesis of PhSH-related diseases.

Two-dimensional design strategy to construct smart dual-responsive fluorescent probe for the precise tracking of ischemic stroke

Two-dimensional design strategy to construct smart dual-responsive fluorescent probe for the precise tracking of ischemic stroke

Construction of a pH- and viscosity-switchable near-infrared fluorescent probe and its imaging application

Viscosity is a parameter used to measure the fluidity of liquids and a key indicator in evaluating the states of body fluid in biological tissues and lesions. Most traditional detection methods have many drawbacks such as a short emission wavelength and interference by background fluorescence. Inspired by the multiple double bond structure of retinal, a novel pH and viscosity dual-response fluorescent probe (Rh-TR) was constructed in this study. Rh-TR exhibited two emission signals centered at 510 and 660 nm. As the pH of the phosphate-buffered saline increased, the fluorescence at 510 nm increased by about 124-fold, while the change in fluorescence at 660 nm was not obvious. When detecting the change in viscosity using the probe, the fluorescence at 510 nm decreased by about 85 %, while the fluorescence at 660 nm increased by over 20-fold. The probe also showed high selectivity and little toxicity. As demonstrated by the biological imaging experiment, the probe successfully imaged changes in the pH and viscosity of cells and in a live animal model of zebrafish. Considering the unique structure of Rh-TR with retinal and its pH- and viscosity-switchable spectral property, the probe may find further application in detecting viscosity-related diseases and industrial detection.

A lysosome-targeted dual-responsive fluorescent probe for simultaneous detection of NO and H2S in living cells, wild-type C. elegans and zebrafish

A lysosome-targeted dual-responsive fluorescent probe YX-N3-OPD was synthesized to detect two major signaling molecules NO and H2S based on nucleophilic aromatic substitution reaction and reduction simultaneously. The photophysical properties revealed that YX-N3-OPD exhibited strong green fluorescence emission at 530 nm in NO and/or H2S. The specific detection and recognition mechanism of NO and/or H2S was verified by high-resolution mass spectrometry. The probe showed significant fluorescence enhancement in cancer cells, especially in glioblastoma cells (9L/lacZ), mainly accumulated in lysosomes. In addition, YX-N3-OPD was also applied for NO and H2S monitoring in wild-type C. elegans and zebrafish with relatively high selectivity and sensitivity.

A dual-emission fluorescent probe for simultaneous detection of singlet oxygen and hypochlorous acid in lipid droplets

Singlet oxygen (1O2) and hypochlorite (ClO-) are two important reactive oxygen species. They coexist in living systems and can be interconverted in organisms, being involved in a variety of pathophysiological processes. Lipid droplets (LDs) are organelles that transport and store lipids, and 1O2 and ClO- tend to destroy unsaturated lipids, leading to lipid oxidation and causing a range of diseases. Here we proposed the first dual-response fluorescent probe DBCC that can simultaneously distinguish 1O2 and ClO- within LDs. The probe is almost non-fluorescent, while significant fluorescence enhancement can be observed in the blue and green channels after treatment with 1O2 and ClO-, respectively. With the excellent properties of high sensitivity, good selectivity and wide tolerance range, the probe DBCC has been successfully used to detect 1O2 and ClO- in HepG2 cells and zebrafish, and faultlessly realized endogenous and exogenous 1O2 and ClO- imaging in LDs. This work provides a powerful analytical tool for further investigating the levels of 1O2 and ClO- in LDs and associated diseases.

Near-infrared dual-response fluorescent probe for detection of N2H4 and intracellular viscosity changes in biological samples and various water samples

N2H4 is a common raw material used in the production of pesticides and has good water solubility, so it may contaminate water sources and eventually enter living organisms, causing serious health problems. Viscosity is an important indicator of the cellular microenvironment and an early warning signal for many diseases. The high reactivity of hydrazine depletes glutathione (GSH) in hepatocytes, causing oxidative stress ultimately leading to significant changes in intracellular viscosity and even death. Therefore, it is particularly important to develop an effective method to detect N2H4 and viscosity in environmental and biological systems. On this basis, we developed two fluorescent probes, BDD and BHD, based on xanthene and 2-benzothiazole acetonitrile. The experimental results show that BHD and BDD have good imaging capabilities for N2H4 in cells, zebrafish and Arabidopsis. BHD and BDD also showed sensitive detection and fluorescence enhancement in the near-infrared region when the intracellular viscosity was changed. Notably, the probe BDD has also successfully imaged N2H4 in a variety of real water samples.

Dual-response fluorescent probe for simultaneously detecting HOCl and Zn2+ in environmental water samples and evaluating the oxidative stress in the zebrafishes induced by pollutants via fluorescence imaging

Both of hypochlorous acid (HOCl) and Zn2+ ions play important roles in fundamental biological processes. They not only simultaneously exist in the biological organisms, but also coexist in the environment. Thus, developing a novel method for simultaneous detection of HOCl and Zn2+ is very necessary. Herein, the nitrogen- and sulfur-codoped carbon dots (N,S-CDs) were synthesized through a simple and rapid microwave method, it emits dual-emission fluorescence centered at 452 nm and 685 nm, which has specific recognition to HOCl and Zn2+, respectively, and they do not interfere with each other. A dual-emission and dual-response fluorescent probe was thus developed for the simultaneous detection of HOCl and Zn2+ in the environment and fluorescence imaging in the zebrafishes. The limits of detection for HOCl and Zn2+ are 27.0 nM and 13.0 nM, respectively. It has been successfully applied for the simultaneous determination of HOCl and Zn2+ in real water samples. The sensing mechanisms of the N,S-CDs to HOCl and Zn2+ were also discussed in detail. Moreover, the fluorescent probe not only can be used to real-time dynamic monitoring of HOCl and Zn2+ in the zebrafishes, but also can be applied to evaluate the oxidative stress in the zebrafishes induced by pollutions via fluorescence imaging. This work provided a new strategy for developing multifunctional fluorescent probes, and a potential application prospect for environmental monitoring and evaluating the health risks of environmental pollutants to organisms.

Simultaneous detection of peroxynitrite and viscosity in diabetes by a dual response fluorescent probe

Diabetes is a common metabolic disease worldwide, which often has serious complications and seriously threatens human health. Abnormal viscosity and peroxynitrite (ONOO−) concentration are closely related to diabetes. Herein, we report a fluorescent dual response probe AO, which can simultaneously detect the changes of viscosity and ONOO− in diabetes at dual fluorescence channels. AO sensitively responds to viscosity and ONOO− with significant emission signal changes at 710 nm and 645 nm, respectively. Moreover, AO selectively targets mitochondria and generates a new compound when detecting ONOO−, which can spontaneously target and light up another type of organelle (lipid droplets), thus showing spatially separated dual-channel fluorescence signals. More notably, AO was successfully used to simultaneously detect the viscosity and ONOO− variations in hyperglycemic cells, diabetes and its complications (using mice models) for the first time, which may lead to a better understanding of the physiological and pathological phenomena of diabetes.

Mitochondria-targeted fluorescent probe for simultaneously imaging viscosity and sulfite in inflammation models.

Many diseases in the human body are related to the overexpression of viscosity and sulfur dioxide. Therefore, it is essential to develop rapid and sensitive fluorescent probes to detect viscosity and sulfur dioxide. In the present work, we developed a dual-response fluorescent probe (ES) for efficient detection of viscosity and sulfur dioxide while targeting mitochondria well. The probe generates intramolecular charge transfer by pushing and pulling the electron-electron system, and the ICT effect is destroyed and the fluorescence quenched upon reaction with sulfite. The rotation of the molecule is inhibited in the high-viscosity system, producing a bright red light. In addition, the probe has good biocompatibility and can be used to detect sulfite in cells, zebrafish and mice, as well as upregulation of viscosity in LPS-induced inflammation models. We expect that the dual response fluorescent probe ES will be able to detect viscosity and sulfite efficiently, providing an effective means of detecting viscosity and sulfite-related diseases.

A dual-color ESIPT-based probe for simultaneous detection of hydrogen sulfide and hydrazine.

Hydrogen sulfide (H2S) and hydrazine (N2H4) are toxic compounds in environmental and living systems, and hydrogen sulfide is also an important signaling molecule. However, in the absence of dual-color probes capable of detecting both H2S and N2H4, the ability to monitor the crosstalk of these substances is restricted. Herein, we developed an ESIPT-based dual-response fluorescent probe (BDM-DNP) for H2S and N2H4 detection via dually responsive sites. The BDM-DNP possessed absorbing strength in the detection of H2S and N2H4, with a large Stokes shift (156 nm for H2S and 108 nm for N2H4), high selectivity and sensitivity, and good biocompatibility. Furthermore, BDM-DNP can be utilized for the detection of hydrogen sulfide and hydrazine in actual soil, and gaseous H2S and N2H4 in environmental systems. Notably, BDM-DNP can detect H2S and N2H4 in living cells for disease diagnosis and treatment evaluation.

Dual-responsive near-infrared turn-on fluorescent probe for cancer stem cell-specific visualization.

Aldehyde dehydrogenase 1A1 (ALDH1A1) stands out as one of the most reliable intracellular biomarkers for stem cells because it is expressed in both cancer stem cells (CSCs) and normal somatic stem cells (NSCs). Although several turn-on fluorescent probes for ALDH1A1 have been developed to visualize CSCs in cancer cells, the discrimination of CSCs from NSCs is difficult. We here report an AND-type dual-responsive fluorescent probe, CHO_βgal, the near-infrared fluorescence of which can be turned on after responding to both ALDH1A1 and β-galactosidase. The AND-type dual responsiveness enables CSCs to be clearly visualized, whereas NSCs are non-emissive in microscopy. CSC-positive metastasis model lungs were successfully discriminated from normal lungs in ex vivo staining experiments using CHO_βgal, whereas the single-input ALDH1A1-responsive probe failed to achieve this discrimination owing to pronounced false-positive fluorescence output from lung NSCs. In tissue slice staining experiments, even in the presence of adjacent normal tissues, the peripheral region-specific localization of CSCs was clear. The versatility of CHO_βgal holds promise not only as a fundamental in vitro research tool for visualizing CSCs but also as a valuable asset in practical tissue staining diagnosis, significantly contributing to the assessment of cancer malignancy.

Dual-channel versatile molecular sensing platform for individual and successive HClO and H2S detection: Applicable in toxic alerts of environmental samples and living organisms

In this study, we have successfully developed a novel dual-response fluorescent probe, NACou, designed for the visual and quantitative detection of HClO/H2S in real water samples and liquid beverages by a thin-film sensing platform. Additionally, NACou demonstrated efficacy for sensing HClO/H2S in HeLa cells, plants and zebrafish through distinct fluorescent channels, yielding satisfactory results. NACou exhibited a multi-modal fluorescence response mechanism for detecting HClO and H2S with remarkable low detection limits of 27.8 nM and 34.4 nM, accompanied by outstanding fluorescent enhancement (209-fold and 148-fold, respectively). These advantages position NACou as a potent molecular tool for HClO and H2S sensing. The specific recognition performance of NACou towards HClO/H2S were confirmed through fluorescence spectroscopy, mass analysis and UV–vis spectroscopy. Importantly, the thin-film sensing platform with the visible fluorescence change can enable rapid assays for water quality and food safety monitoring, showcasing significant practical application value. Impressively, NACou has been employed in warning against liver injury induced by multiple drugs, allowing for the exploration of the pathogenesis and degree of drug-induced injury.