Dual-channel Fluorescent Probe Research Articles

Dual-channel discrimination of two lethal chemical warfare agents using an ESIPT-ICT-based fluorescent probe

Among all kinds of chemical warfare agents, only cyanide and nerve agents can cause massive mortality at low concentrations. In this work, a dual-channel fluorescent probe CWAs-Thia capable of detecting cyanide and nerve agents is presented. The two reactive recognition units, pyridine and the thiazole-2-carbonyl group, of the probe for cyanide and nerve agents, respectively, produced red and blue fluorescent responses, respectively, which were attributed to excited-state intramolecular proton transfer and intramolecular charge transfer. CWAs-Thia is the first probe that can selectively recognize cyanide and nerve agent. And it has proven to be effective in visualizing cyanide and nerve agents in living cells.

A dual-channel fluorescent probe targeting lysosomes for differential detection of Cys/Hcy and GSH: Applications in food, pharmaceutical analysis and bioimaging

Thiols function as antioxidants in food, prolonging shelf life and enhancing flavor. Moreover, thiols are vital biomolecules involved in enzyme activity, cellular signal transduction, and protein folding among critical biological processes. In this paper, the fluorescent probe PYL-NBD was designed and synthesized, which utilized the fluorescent molecule pyrazoline, the lysosome-targeted morpholine moiety, and the sensing moiety NBD. Probe PYL-NBD was tailored for the recognition of biothiols through single-wavelength excitation, yielding distinct fluorescence emission signals: blue for Cys, Hcy, and GSH; green for Cys, Hcy. Probe PYL-NBD exhibited rapid reaction kinetics (<10 min), distinct fluorescence response signals, and low detection limits (15.7 nM for Cys, 14.4 nM for Hcy, and 12.6 nM for GSH). Probe PYL-NBD enabled quantitative determination of Cys content in food samples and L-cysteine capsules. Furthermore, probe PYL-NBD had been successfully applied for confocal imaging with dual-channel detection of biothiols in various biological specimens, including HeLa cells, zebrafish, tumor sections, and Arabidopsis thaliana.

Dual-channel fluorescent probe utilizing hydrazone for Zn2+ and Hg2+ detection: Exploring distinct signaling mechanisms and applications in bioimaging and latent fingerprint analysis

A facile Schiff base probe C1 has been developed and evaluated using 1H, 13C NMR, and mass spectroscopy. Fluorescence investigations were conducted to determine probe-sensing behavior. Probe C1 demonstrated sensitive, and selective recognition of Zn2+ and Hg2+ ions by using two distinct processes (i.e., chelation enhanced fluorescence effect and fluorescence quenching) over other cations in ACN-H2O (8:2, v/v). The Zn2+ and Hg2+ detection limits of 0.042 and 0.081 μM are achieved with the probe C1. C1 has a binding constant of 3.33 × 103 M−1 for Zn2+ and an emission quenching constant of 1.22 × 104 M−1 for Hg2+. A Jobs plot was used to determine the binding stoichiometry of C1 and Zn2+/Hg2+ ions ratio is 1:1. By using Zn2+/Hg2+ and EDTA additions alternately, the probe's reusability was investigated. Moreover, the probe was found to be highly cytocompatibility using an MTT assay in a mouse fibroblast cell line (NIH3T3). Further, the probe was found to visualize Zn2+ and Hg2+ ions inside the cells, indicating the potential application of the probe as an intracellular probe to detect Zn2+ and Hg2+ ions. The probe shows very good solid-state emission behavior, this motivates us to develop latent fingerprint detection and anticounterfeiting applications.

Water-Soluble Dual-Channel Fluorescent Probe for Sensitive Detection of Biothiols In Vitro and In Vivo.

Benefiting from high spatiotemporal resolution, deep tissue penetration, and excellent sensitivity, fluorescence imaging technology has been widely applied in cancer diagnosis and treatment. In recent years, a large number of fluorescent probes for monitoring the levels of endogenous biothiols have been reported, which have significant implications for cancer diagnosis and treatment. However, most probes still suffer from poor biological compatibility and easy attachment by the environment. This work presents the development of a water-soluble dual-channel fluorescent probe, named MAL-NBD, for sensitively detecting biothiols. Nonfluorescent MAL-NBD is transformed into fluorescent groups MAL and NBD-SR/NR through nucleophilic substitution by biologically active thiols, producing dual-channel fluorescence signals for precise detection of biologically active thiols. Taking advantage of the excellent biocompatibility and low biotoxicity, MAL-NBD is successfully used for imaging HeLa cancer cells and zebrafish larvae, promoting its potential application for the precise detection of biological thiols involved in physiological and pathological processes.

Dual-channel fluorescent probe for discriminative detection of H2S and N2H4: Exploring sensing mechanism and real-time applications

A highly efficient system incorporates the real-time visualization of the two toxic molecules (H2S and N2H4) and the recognition of corresponding transforms using a fluorescent sensor. In this paper, a dual-responsive probe (QS-DNP) based on methylquinolinium-salicyaldehyde-2,4-dinitrophenyl was developed that can simultaneously detect H2S and N2H4 at two independent fluorescent channels without signal crosstalk. QS-DNP showed excellent anti-interference, high selectivity, outstanding water solubility, low LOD values (H2S: 51 nM; N2H4: 40 nM), low cytotoxicity, and mitochondrial localization properties. The 2,4-dinitrophenyl site was sensitive to H2S, and the CC bridge was reactive to N2H4, with strong fluorescence at 680 and 488 nm, respectively. The wavelength gap between these two channels is 192 nm; verify that there is no signal crosstalk throughout detection. By this means, the probe was used to simultaneously detect H2S and N2H4 in real soil samples, food samples, and living cells. The endogenous H2S and N2H4 were monitored in HeLa cells and investigated the mitochondria organelle of living cells with a positive charge on QS-DNP. Overall, all results emphasize that the QS-DNP probe is a powerful tool for the simultaneous detection of H2S and N2H4 and presents a potential new sensing approach.

Dual-channel fluorescent probe for simultaneously detecting H2S and viscosity/polarity and its application in non-alcoholic fatty liver, tumor tissue, and food spoilage

Intracellular hydrogen sulfide (H2S) and micro-environments (e.g. viscosity and polarity) are tightly correlated with various physiological/pathological processes, making them potential biomarkers of many diseases. However, the simultaneously detecting H2S, viscosity, and polarity in inflammation, non-alcoholic fatty liver (NAFL) and tumor tissues containing clinical cancer patient samples has not been achieved yet due to the lack of effective tools. Herein, we presented a mitochondria-targeting near-infrared (NIR) fluorescent probe MQA-DNP) for simultaneously monitoring H2S, viscosity/polarity through dual channels. The probe could specifically recognize H2S via far-red emission (λem = 634 nm), and exhibited high sensitivity toward micro-environmental viscosity/polarity in the NIR channel (λem > 714 nm). Facilitated with the probe, we revealed for the first time that inflammation cells and NAFL tissues are accompanied by an up-regulation of H2S, as well as an increase in viscosity level (and/or decrease in polarity degree). Surprisingly, the simultaneous visualization of H2S, viscosity/polarity has also been achieved not only at the cancer cells and tumor models, but also in clinical cancer patients. Compared to detecting a single biomarker, the simultaneous tracking multi-markers through multi-channels seems to be more reliable for visual medical diagnosis of mitochondria-related diseases. Furthermore, inspired by the significant color changes and high sensitivity of MQA-DNP reaction with H2S, MQA-DNP has been successfully used to detect H2S released during food spoilage through test strips, and has great potential for application in complex environments systems.

A two-pronged detection of atherosclerosis with a dual-channel fluorescent probe for viscosity and hypochlorous acid

The precise detection of atherosclerosis has been a research hot point since its importance in the diagnosis of cardiovascular diseases. Here we synthesized a near-infrared fluorescent probe AS-CN with a D − π − A structure, enabling independent dual-response determination of hypochlorous acid (HOCl) and viscosity during AS progression. The performance of this probe is excellent, with large Stokes shift, low detection limit, high sensitivity, and good biocompatibility. The simultaneous variations of HOCl and viscosity during the formation of foam cells were successfully investigated for the first time. Especially, we further applied it to achieve the non-invasive in vivo detection in the atherosclerotic mouse and realized the evaluation of atherosclerosis from chemical (ROS) and physical (viscosity) dimensions, respectively. We hope that AS-CN acts as a potent tool in the detection of atherosclerosis.

H2S-activated fluorescent probe enables dual-channel fluorescence tracking of drug release in tumor cells

Nowadays, the selective release of therapeutic drugs into tumor cells has become an important way of tumor treatment due to the high side effects of chemotherapy drugs. As one of the gas mediators, hydrogen sulfide (H2S) is closely related to cancer. Due to the high content of H2S in tumor cells, it can be used as a signaling molecule that triggers the release of drugs to achieve the selective release of therapeutic drugs. In addition, dual-channel fluorescence imaging technology can be better applied to monitor the drug delivery process and distinguish the state before and after drug release, so as to better track the effect of drug therapy. Based on this, we used NBD amines (NBD-NHR) as the recognition group of H2S and connected the tyrosine kinase inhibitor crizotinib to construct an activated dual-channel fluorescent probe CZ-NBD. After the probe enters the tumor cells, it consumes H2S and releases crizotinib, which is highly toxic to the tumor cells. Importantly, the probe displays significant fluorescence changes in different cells, enabling not only the screening of tumor cells, but also tracking and monitoring drug release and tumor cell activity. Therefore, the construction of probe CZ-NBD provides a new strategy for drug release monitoring in tumor cells.

Rational Design of a Dual-Channel Fluorescent Probe for the Simultaneous Imaging of Hypochlorous Acid and Peroxynitrite in Living Organisms.

Hypochlorous acid (HOCl) and peroxynitrite (ONOO-) are two important highly reactive oxygen/nitrogen species, which commonly coexist in biosystems and play pivotal roles in many physiological and pathological processes. To investigate their function and correlations, it is urgently needed to construct chemical tools that can track the production of HOCl and ONOO- in biological systems with distinct fluorescence signals. Here, we found that the coumarin fluorescence of coumarin-benzopyrylium (CB) hydrazides (spirocyclic form) is dim, and their fluorescence properties are controlled by their benzopyran moiety via an intramolecular photo-induced electron transfer (PET) process. Based on this mechanism, we report the development of a fluorescent probe CB2-H for the simultaneous detection of HOCl and ONOO-. ONOO- can selectively oxidize the hydrazide group of CB2-H to afford the parent dye CB2 (Absmax/Emmax = 631/669 nm). In the case of HOCl, it undergoes an electrophilic attack on the benzopyran moiety of CB2-H to give a chlorinated product CB2-H-Cl, which inhibits the PET process within the probe and thus affords a turn-on fluorescence response at the coumarin channel (Absmax/Emmax = 407/468 nm). Due to the marked differences in absorption/emission wavelengths between the HOCl and ONOO- products, CB2-H enables the concurrent detection of HOCl and ONOO- at two independent channels without spectral cross-interference. CB2-H has been applied for dual-channel fluorescence imaging of endogenously produced HOCl and ONOO- in living cells and zebrafish under different stimulants. The present probe provides a useful tool for further exploring the distribution and correlation of HOCl and ONOO- in more biosystems.

Dual-Channel Fluorescent Probe for Detecting Viscosity and ONOO- without Signal Crosstalk in Nonalcoholic Fatty Liver.

Nonalcoholic fatty liver disease (NAFLD) is a global health issue. Peroxynitrite and liver viscosity have recently been found to be potential biomarkers of NAFLD. Therefore, it is of great significance to develop dual-response fluorescent probes for simultaneous detecting peroxynitrite and viscosity. We report herein a new probe (CQ) that can simultaneously detect peroxynitrite and viscosity at two independent fluorescent channels without signal crosstalk. CQ shows high selectivity, rapid response, good water solubility, low cytotoxicity, and mitochondrial localization properties. In particular, CQ responds sensitively to viscosity and peroxynitrite with off-on fluorescence changes at 710 and 505 nm, respectively. The wavelength gap between these two channels is more than 200 nm, ensuring that there is no signal crosstalk during detection. With this property, the probe was applied to simultaneously detect mitochondrial viscosity and peroxynitrite and image the changes of liver viscosity and peroxynitrite concentration during the pathogenesis of NAFLD. All results show that the CQ probe is a powerful tool for simultaneous detection of viscosity and peroxynitrite and provides a potential new diagnostic method for NAFLD.

Dual-Channel Fluorescent Probe for the Detection of Peroxynitrite and Glutathione in Mitochondria: Accurate Discrimination of Inflammatory and Progressing Tumor Cells.

Distinguishing between normal, inflammatory, and progressing tumor cells plays a vital role in early diagnoses and clinical studies. The simultaneous quantification of multiple biomarkers in cells can reveal cellular heterogeneity, which contributes to the discrimination of different types of cells. Herein, a dual-channel fluorescent probe has been developed for monitoring peroxynitrite (ONOO-) and glutathione (GSH) to accurately discriminate normal cells, inflammatory cells, and progressing cancer cells. The probe can monitor exogenous and endogenous mitochondrial GSH and ONOO- in living cells and zebrafish by green (530 nm, G530) and red (630 nm, R630) emission based on its good selectivity and low biotoxicity. GSH and ONOO- are visualized via fluorescence imaging, and the corresponding output signals can be employed to differentiate nontumorigenic, malignant, and metastatic breast cells in cocultured cells. Furthermore, the accurate discrimination among normal, inflammatory, and cancerous cells is achieved through the changes in the dual-channel fluorescence signal, which shows great potential for the diagnosis of inflammation and cancer diseases.

A stilbene derivative as dual-channel fluorescent probe for mitochondrial G-quadruplex DNA in living cells

G-quadruplex DNA has attracted the widespread attention as a novel target of anticancer strategy. Herein, two novel stilbene derivatives 2a and 2b were designed and synthesized under mild reaction conditions, and their interactions with G-quadruplex DNA, cytotoxicity, and distribution in living cells were investigated in detail. Both compounds display a low cytotoxicity and the higher affinity to G-quadruplex DNA than to the other secondary structures, including duplex, single-stranded and i-motif DNA, moreover, the affinity of 2b with m-allyl pyridine salt group to G-quadruplex DNA is about 10-fold stronger than that of 2a with p-allyl pyridine salt group. The interactions of the compounds with the promoter G-quadruplexes are enthalpy-driven by an ITC assay. 2a and 2b not only stabilize the G-quadruplex structure but also induce the G-rich sequences (bcl-2, HRCC and KSS) to fold into the mixed-type G-quadruplex in Na+/K+ free Tris-HCl buffer at pH 7.0, and 2b presents the higher stabilization to G-quadruplex than 2a by a FRET-melting assay. 2b presents a dual-emission at 508 and 600 nm and gives a turn-on and stronger and more sensitive fluorescence response over 2a to the promoter (bcl-2, c-kit 2 and c-myc) and mitochondrial (HRCC and KSS) G-quadruplex DNA at both emission wavelengths, moreover, the peak at 508 nm is blue-shifted to 466 nm after binding to DNA. The blue and red dual-channel CLSM images indicate that 2b is mainly distributed in the mitochondrion of living HepG2 cells. The results show that 2b is a potential dual-channel fluorescent probe for mitochondrial G-quadruplex DNA in living cells.

A novel dual-channel fluorescent probe for selectively and sensitively imaging endogenous nitric oxide in living cells and zebrafish

Nitric oxide (NO) plays various physiological and pathological roles in lots of biological processes. It is crucial to detect NO sensitively and selectively in vivo and in vitro as homeostasis of NO is closely related to various diseases. Herein, a novel dual-channel fluorescent dye (ENNH2) based on dicarboxyimide anthracene was developed as a highly sensitive and selective probe to detect NO in living systems using the dual-channel fluorescence. ENNH2 can emit bright red fluorescence due to the intramolecular charge transfer (ICT) from the amino group at the 6-position of 1,2-dicarboxyimide anthracene to the conjugated aromatic ring, and the ICT is effectively inhibited by the reductive deamination of the amino in the presence of NO to obtain the remarkable strong green emission with the excellent sensitivity (5.52 nM). Promisingly, ENNH2 exhibits an excellent performance in endogenous NO dual-channel fluorescence imaging of RAW 264.7 cells and zebrafish.

A dual-channel fluorescent ratio probe with hypoxia targeting and hypoxia activation capacity for tumour imaging

Illustration of the fluorescent ratio probe (OPDEA/BOD-NO2) with hypoxia targeting and hypoxia activation capacity for in vivo tumour imaging.

Dual-Channel Fluorescent Probe for the Simultaneous Monitoring of Peroxynitrite and Adenosine-5'-triphosphate in Cellular Applications.

Changes in adenosine triphosphate (ATP) and peroxynitrite (ONOO–) concentrations have been correlated in a number of diseases including ischemia-reperfusion injury and drug-induced liver injury. Herein, we report the development of a fluorescent probe ATP-LW, which enables the simultaneous detection of ONOO– and ATP. ONOO– selectively oxidizes the boronate pinacol ester of ATP-LW to afford the fluorescent 4-hydroxy-1,8-naphthalimide product NA-OH (λex = 450 nm, λem = 562 nm or λex = 488 nm, λem = 568 nm). In contrast, the binding of ATP to ATP-LW induces the spirolactam ring opening of rhodamine to afford a highly emissive product (λex = 520 nm, λem = 587 nm). Due to the differences in emission between the ONOO– and ATP products, ATP-LW allows ONOO– levels to be monitored in the green channel (λex = 488 nm, λem = 500–575 nm) and ATP concentrations in the red channel (λex = 514 nm, λem = 575–650 nm). The use of ATP-LW as a combined ONOO– and ATP probe was demonstrated using hepatocytes (HL-7702 cells) in cellular imaging experiments. Treatment of HL-7702 cells with oligomycin A (an inhibitor of ATP synthase) resulted in a reduction of signal intensity in the red channel and an increase in that of the green channel as expected for a reduction in ATP concentrations. Similar fluorescence changes were seen in the presence of SIN-1 (an exogenous ONOO– donor).

Simultaneous imaging of hypochlorous acid and nitric oxide in live cells based on a dual-channel fluorescent probe

Both reactive oxygen species (ROS) and reactive nitrogen species (RNS) are inevitably produced during normal human metabolism. Various ROS and RNS together form tangled networks that play important roles in many physiological and pathological processes. Here we used 1,8-naphthalene diamine as a reactive group to develop a fluorescent probe, N-[2-(6-phenylethynyl)quinolinylmethyl]-1,8-diamino naphthalene (QBN), for HOCl and NO. QBN showed a “turn-on” fluorescent response at 464 nm to HOCl in the range of 0–75 μM with rapid responding time (10 s) and detection limit (0.11 ± 0.03 μM). Furthermore, a “turn-on” fluorescent responses at 512 nm to NO in the range of 0–40 μM with responding time (20 s) and detection limit (25.7 ± 3.4 nM) was found. The response mechanisms of QBN to HOCl and NO were discussed based on mass analysis of the different products. The dual-channel probe was then successfully applied for simultaneous imaging of both exogenous and endogenous HOCl and NO in live cells.

A dual channel fluorescent probe with pH-based specificity to lysosomes for multicolor imaging and localization

We herein innovatively constructed the pure pH-dual channel probe DNOXA by the reaction of oxalyl chloride with 4-dimethylaminobenzaldehyde. DNOXA has two response sites for acidity, which are corresponding to pH-dual channel responses. When adding acid to pH 5.0, the protonation in the response site I enhanced the push-pull effect, and the absorption maxima of the optical spectra bathochromically shifted from 463 to 652 nm. When sequentially adding acid to pH 1.6, the response site II is awakened to work. The enhanced push-pull system was interrupted, and the absorption maxima hypochromatically shifted from 652 to 506 nm. In this process, the ratiometric change in the probe can be observed by naked-eye. Furthermore, quantum-chemical study well explained and supported the difference of their maxima absorption. Based on the excellent response of DNOXA to pH in vitro, DNOXA had commendable specific imaging and localization ability for lysosomes, undergoing dual channel images of protonation at the acidic microenvironment.

Individual and successive detection of H2S and HClO in living cells and zebrafish by a dual-channel fluorescent probe with longer emission wavelength

Reactive oxygen species (ROS) and reactive sulfur species (RSS) participate in many physiological activities and help maintaining the redox homeostasis in biological system. The complicated intrinsic connection between specific ROS/RSS needs to be further explored. Herein, a novel fluorescent probe (MB-NAP-N3) with longer emission wavelength has been rationally designed and synthesized based on the conjugation of the methylene blue moiety and the naphthalimide moiety for the detection of hypochlorous acid (HClO) and hydrogen sulfide (H2S). The dual-signal probe exhibits rapid turn-on fluorescence responses for individual and successive detection of H2S and HClO in green and red channels, respectively. Owning to its advantages such as fast response, good selectivity and high sensitivity, the probe was successfully applied to detect endogenous and exogenous HClO/H2S in living cells. Furthermore, the outstanding luminescence performance makes it suitable for the visualization of the in vivo interaction between the two analytes in zebrafish.

A Highly Selective Turn-on Fluorescent and Naked-eye Colourimetric Dual-channel Probe for Cyanide Anions Detection in Water Samples.

A highly selective turn-on fluorescent and naked-eye colourimetric dual-channel probe for cyanide anions (CN-) has been designed and characterized. In the mixed solution (DMSO / H2O, 9:1, v / v), only CN- could cause an increase in the UV absorption intensity and the corresponding fluorescence intensity increased, and other anions had no significant effect on the probe. After treatment with cyanide in the probe solution, the solution showed a noticeable colour change, from light yellow to purple. Moreover, a fluorescence spectrophotometer can be used to observe that the fluorescence intensity of the solution is significantly enhanced. The response of the colourimetric and fluorescent dual-channel probe to CN- was attributed to nucleophilic addition, and the mechanism was determined by 1H NMR spectroscopy. In addition, this probe was used to detect CN- in actual water samples, including river water, drinking water, and tap water. The spiked CN- recovery rate is very high (97.2%-100.06%), and analytical precision is also very high (RSD < 2%), which shows its feasibility and reliability for detecting cyanide ions in actual water samples.

A dual-channel fluorescent probe for monitoring pH changes in lysosomes during autophagy

A new fluorescent probe RD for visualizing starvation-induced autophagy in living cells and for distinguishing between neutral and acidic lysosomes in the process of autophagy.