Design Of Fluorescent Probes Research Articles

Molecular design and application of fluorescent probes for drug discovery research

Molecular design and application of fluorescent probes for drug discovery research

The investigation of fluorescence and metal interaction properties of racemic 7,8,9,10-tetrahydro-3-hydroxy-4-(1-hydroxyethyl)benzo[c]chromen-6-one

The design or investigation of fluorescence probes continues to receive attention with respect to the diverse applications of spectrofluorometry. Depending on the highly sensitive character, fluorescence spectroscopy-based methodologies have been widely used in recent years in different sciences, including analytical, environmental, and medicinal chemistry areas. In our previous works, we have shown the iron (III) selective on-off sensor properties of benzo[c]chromen-6-one derivatives. In this study, we have extrapolated this research to 4-substituted analogues and investigated both fluorescent and metal interaction properties. Following the synthesis and structure identification studies, (±)-7,8,9,10-tetrahydro-3-hydroxy-4-(1-hydroxyethyl)benzo[c]chromen-6-one was found as a fluorescent molecule displaying fluorescence enhancement in the presence of metals. This feature has been found quite different in comparison to the previous urolithins investigated. This finding suggested the substituent dependent effects and variations on the fluorescent properties of benzo[c]chromen-6-one system.

Design of Nir-Ii High Performance Organic Small Molecule Fluorescent Probes and Summary of Their Biomedical Applications

Design of Nir-Ii High Performance Organic Small Molecule Fluorescent Probes and Summary of Their Biomedical Applications

Current Probes for Imaging Carbonylation in Cellular Systems and Their Relevance to Progression of Diseases.

Oxidative stress resulted from reactive oxygen or nitrogen species in biological systems has a significant role in the diagnosis/progression of several human diseases. Human diseases associated with oxidative stress include Alzheimer's disease, chronic lung disease, chronic renal failure, cancer, diabetes, and fibrosis. In oxidative stress conditions, carbonylation process can be described as one of the most common modifications in biomolecules that takes place in the presence of carbonyl (C = O) groups which are introduced into molecules by direct metal-catalyzed oxidation of certain amino acids or indirectly by reaction with the oxidation of lipids and sugars. At a molecular cellular level, carbonylation can cause some defective biological consequences or chemical transformations in cells. During this process, specifically, carbonylated proteins can be accumulated in cells and trigger to develop some diseases in human body. The role of the accumulation of carbonylated proteins in the progression of several diseases has also been reported in the literature, such as neurodegenerative diseases, diabetes, obesity, aging, and cancer. Early detection of carbonylation process is, therefore, very critical to monitor these diseases at an early stage. Finding a suitable biomarker or probe is very challenging due to the need for multiple criteria: high fluorescence efficiency, stability, toxicity, and permeability. If they are designed with a good strategy, these probes are highly effective in cell biology applications and they can be used as good diagnostic tools for monitoring oxidative stress-induced carbonylation in relevant diseases. This review highlights the design and use of recent fluorescent probes for visualization of carbonylation in cellular systems and the relationship between oxidative stress and carbonyl species for causing long-term disease complications.

Rational Design of a Symmetric Double Rotor-Based Fluorescent Probe for Revealing Lds Viscosity Changes During Ferroptosis

Rational Design of a Symmetric Double Rotor-Based Fluorescent Probe for Revealing Lds Viscosity Changes During Ferroptosis

Design, synthesis and evaluation of liver-targeting fluorescent probes for detecting mercury ions.

Three fluorescent glycosyl-rhodamine probes with good selectivity and sensitivity toward Hg2+ were developed. The detection limit of the probes toward Hg2+ is as low as 94.6 nM, which can be used to detect trace Hg2+ in solution. 1 : 1 stoichiometry was the most possible recognition mode of the probes toward Hg2+, and the OFF/ON mechanism of the probes toward Hg2+ could be attributed to the closing or opening of the rhodamine spiral structure caused by Hg2+. The detection of Hg2+ is reversible, which is beneficial for the recycling of probes. Moreover, these low cytotoxic probes can be safely and selectively applied to monitor Hg2+ levels in hepatocytes, and the fluorescence response follows a trend of Rho-Gal > Rho-Lac > Rho-Glu in HepG2 cells because the galactose group in Rho-Gal can selectively recognize ASGPR overexpressed on HepG2 cells.

Rational Design of ICT-Based Fluorescent Probe with AIE and NIR Properties for Hypochlorite Determination

In recent years, the easily synthesized fluorescent probes with good photophysical and sensing properties have attracted widespread attention. Herein, by utilizing the fluorescence regulation effect of electron push-pull effect and the oxidation property of hypochlorite (ClO−) to C=N double bonds, we rationally proposed two intramolecular charge transfer (ICT)-based fluorescent probes with typical aggregation-induced emission (AIE) properties for ClO− detection. The synthesis process of the two probes is very convenient, and both of them can exhibit significant colorimetric and fluorescence changes within 3 min in the presence of ClO−. Moreover, compared with Probe A, the Probe B with near-infrared (NIR) fluorescence centered at 677 nm was successfully applied to ClO− determination in tap water and food samples as well as live cell imaging.

Highly Selective Fluorescent Probe Design for Visualizing Hepatic Hydrogen Sulfide in the Pathological Progression of Nonalcoholic Fatty Liver.

Hydrogen sulfide (H2S), emerging as an important gaseous signal, has attracted more and more attention for its key role in chronic fatty liver diseases. However, lacking tools for H2S-specific in situ detection, the changes of endogenous hepatic H2S levels in the pathological progression of chronic liver diseases are still unclear. To this end, we adopted a strategy of combining molecular probe design and nanofunctionalization to develop a highly selective near-infrared (NIR) fluorescent probe, which allows in vivo real-time monitoring of hepatic H2S levels in the process of nonalcoholic fatty liver disease (NAFLD). As a proof of strategy demonstration, we first designed NIR molecular probes for H2S sensing through chemical design and probe screening and then loaded molecular probes into mesoporous silicon nanomaterials (MSNs) with surface encapsulation using poly(ethylene glycol) to construct a highly selective probe MSN@CSN@PEG, with significantly improved selectivity and photostability. Moreover, MSN@CSN@PEG exhibited high selectivity and sensitivity for endogenous H2S in cells and tumors in vivo, eliminating the interference of a high concentration of biothiols and sulfhydryl proteins. Furthermore, the probe was applied to in situ intravital imaging and systematic assessment of hepatic H2S levels in different stages of NAFLD for the first time, which may offer a promising tool for the future study of fatty liver diseases and other chronic liver diseases.

Sensitive Immunofluorescent Detection of the PRAME Antigen Using a Practical Antibody Conjugation Approach.

Bioconjugation of antibodies with various payloads has diverse applications across various fields, including drug delivery and targeted imaging techniques. Fluorescent immunoconjugates provide a promising tool for cancer diagnostics due to their high brightness, specificity, stability and target affinity. Fluorescent antibodies are widely used in flow cytometry for fast and sensitive identification and collection of cells expressing the target surface antigen. Nonetheless, current approaches to fluorescent labeling of antibodies most often use random modification, along with a few rather sophisticated site-specific techniques. The aim of our work was to develop a procedure for fluorescent labeling of immunoglobulin G via periodate oxidation of antibody glycans, followed by oxime ligation with fluorescent oxyamines. Here, we report a novel technique based on an in situ oxime ligation of ethoxyethylidene-protected aminooxy compounds with oxidized antibody glycans. The approach is suitable for easy modification of any immunoglobulin G, while ensuring that antigen-binding domains remain intact, thus revealing various possibilities for fluorescent probe design. The technique was used to label an antibody to PRAME, a cancer-testis protein overexpressed in a number of cancers. A 6H8 monoclonal antibody to the PRAME protein was directly modified with protected-oxyamine derivatives of fluorescein-type dyes (FAM, Alexa488, BDP-FL); the stoichiometry of the resulting conjugates was characterized spectroscopically. The immunofluorescent conjugates obtained were applied to the analysis of bone marrow samples from patients with oncohematological diseases and demonstrated high efficiency in flow cytometry quantification. The approach can be applied for the development of various immunofluorescent probes for detection of diagnostic and prognostic markers, which can be useful in anticancer therapy.

Indole-incorporated-benzoeindolium as a novel mitochondrial and ratiometric fluorescent probe for real-time tracking of SO2 derivatives in vivo and herb samples

Sulfur dioxide (SO2) has been regarded as a well-known environmental pollutant that can cause allergic respiratory responses and even serious diseases, whereas endogenous SO2 as an important gaseous signaling molecule is involved in many physiological and pathological processes. In addition, SO2 and its derivatives (bisulfite and sulfite) are also widely-used preservatives and antioxidants in various foods, beverages and pharmaceuticals, whose concentration level is strictly limited due to the adverse effect on community health. Thus, dynamical determination of SO2 and its derivatives both in vitro and in vivo is highly desirable. Herein, we reported a novel mitochondrial fluorescent probe indole-incorporated-benzoeindolium (InB) for SO2 derivatives with high sensitivity and selectivity in DMSO-H2O (2:3, v/v, 10 mM PBS, pH = 7.40) media. The probe exhibited low detection limit (4.39 nM) and rapidly responsive (300 s) recognition for SO2 derivatives in a ratiometric manner. The sensing mechanism could be attributed to the nucleophilic addition of the electron deficit carbon-carbon double bonds from the linker with HSO3−, resulting in inhibition of the ICT process of the probe after reaction, which was confirmed by high-resolution mass spectroscopy (HR-MS) and 1H NMR techniques. Moreover, the probe was successfully applied to detect HSO3− in Dioscotea opposita Thunb and Gastrodia elata Bl, which represents one of the only two examples of using synthetic fluorescent probes to monitor SO2 derivatives in herb samples to date. Furthermore, its applications in real-time tracking and bioimaging of SO2 derivatives in living cells, zebrafish and nude mice were also achieved. Hopefully, this work provides a promising methodology for the design of other fluorescent probes for SO2 derivatives with tailored performance.

An Edaravone-Guided Design of a Rhodamine-Based Turn-on Fluorescent Probe for Detecting Hydroxyl Radicals in Living Systems.

The hydroxyl radical (·OH), one of the reactive oxygen species (ROS) in biosystems, is found to be involved in many physiological and pathological processes. However, specifically detecting endogenous ·OH remains an outstanding challenge owing to the high reactivity and short lifetime of this radical. Herein, inspired by the scavenging mechanism of a neuroprotective drug edaravone toward ·OH, we developed a new ·OH-specific fluorescent probe RH-EDA. RH-EDA is a hybrid of rhodamine and edaravone and exploits a ·OH-specific 3-methyl-pyrazolone moiety to control its fluorescence behavior. RH-EDA itself is almost nonfluorescent in physiological conditions, which was attributed to the formation of a twisted intramolecular charge transfer (TICT) state upon photoexcitation and the acylation of its rhodamine nitrogen at the 3' position. However, upon a treatment with ·OH, its edaravone subunit was converted to the corresponding 2-oxo-3-(phenylhydrazono)-butanoic acid (OPB) derivative (to afford RH-OPB), thus leading to a significant fluorescence increase (ca. 195-fold). RH-EDA shows a high sensitivity and selectivity to ·OH without interference from other ROS. RH-EDA has been utilized for imaging endogenous ·OH production in living cells and zebrafishes under different stimuli. Moreover, RH-EDA allows a high-contrast discrimination of cancer cells from normal ones by monitoring their different ·OH levels upon stimulation with β-Lapachone (β-Lap), an effective ROS-generating anticancer therapeutic agent. The present study provides a promising methodology for the construction of probes through a drug-guided approach.

Strategic Engineering of Sub‐5 nm Dyes@CDs Nanoassemblies Platform for Super Resolution Imaging

AbstractThe design and construction of fluorescence probes with small size, high quantum yield, photostability, and solubility in an aqueous solution to achieve biomarker have become a focus of interest in super‐resolution bioimaging application. Herein, four kinds of GFP‐like supramolecular nanoassemblies (SNAs) are constructed in a very facile manner by the host–guest interaction between isopercolic acid derivatives (IADs) and cyclodextrin (CD). 1H NMR, Fourier transform infrared spectroscopy (FT‐IR) and single crystal structure well prove the formation of SNAs. Moreover, compared to the initial state, the fluorescence quantum yield of the SNAs is dramatically improved through suppressing nonradiative relaxation pathways and shielding quenchers. The bimolecular exciton–exciton annihilation (bmEA) process under the high pumping fluence has been suppressed effectively. Both the improved gain and the reduced loss facilitate the buildup of stimulated emission and enable the high stimulated emission efficiency of these SNAs. Sub‐5 nm IADs@β‐CDs‐mediated STED imaging exhibits excellent stimulated emission depletion (STED) performance: high photostability, low saturation intensity (≈0.50 MW cm−2), and high resolution (30 nm) under PSTED‐775 nm of 109 MW cm−2. This supramolecular strategy avoids the tedious molecular synthesis process and paves a new pathway for insights into building up novel high‐performance STED probes.

Research Progress of Small Molecule Fluorescent Probes for Detecting Hypochlorite.

Hypochlorous acid (HOCl) generates from the reaction between hydrogen peroxide and chloride ions via myeloperoxidase (MPO)-mediated in vivo. As very important reactive oxygen species (ROS), hypochlorous acid (HOCl)/hypochlorite (OCl−) play a crucial role in a variety of physiological and pathological processes. However, excessive or misplaced production of HOCl/OCl− can cause variety of tissue damage and human diseases. Therefore, rapid, sensitive, and selective detection of OCl− is very important. In recent years, the fluorescent probe method for detecting hypochlorous acid has been developed rapidly due to its simple operation, low toxicity, high sensitivity, and high selectivity. In this review, the progress of recently discovered fluorescent probes for the detection of hypochlorous acid was summarized with the aim to provide useful information for further design of better fluorescent probes.

Illuminating Protein Phase Separation: Reviewing Aggregation-Induced Emission, Fluorescent Molecular Rotor and Solvatochromic Fluorophore Based Probes.

Protein phase separation process involving protein unfolding, misfolding, condensation and aggregation etc. has been associated with numerous human degenerative diseases. The complexity in protein conformational transitions results in multi-step and multi-species biochemical pathways upon protein phase separation. Recent progresses in designing novel fluorescent probes have unraveled the enriched details of phase separated proteins and provided mechanistic insights towards disease pathology. In this review, we summarized the design and characterizations of fluorescent probes that selectively illuminated proteins at different phase separated states with a focus on aggregation-induced emission probes, fluorescent molecular rotors, and solvatochromic fluorophores. Inspired by these pioneering works, a design blueprint was proposed to further develop fluorescent probes that can potentially shed light on the unresolved protein phase separated states in the future.

(INVITED) Fluorescent probes for optical investigation of the plasma membrane

The biological functions of the cell membrane are regulated by its physicochemical properties, such as the transmembrane potential, phase state and hydration. Therefore, the in situ investigation of these properties is attracting a rising interest in the field of membrane biophysics. In these regards, a wide variety of fluorescent membrane probes has been developed, providing powerful tools for fluorescence microscopy and/or fluorescence spectroscopy to study the microscopic analogs of viscosity, polarity, and hydration, as well as the molecular order and electrostatic potential at the sites of their location. Despite the success of this approach, further developments rely on the design of fluorescence probes with optimized photophysical properties. This review discusses the most successful fluorescent molecular probes and their application to the study of the plasma membrane viscosity, lipid order and potential.

Ratiometric Imaging of MMP-2 Activity Facilitates Tumor Detection Using Activatable Near-Infrared Fluorescent Semiconducting Polymer Nanoparticles.

Enzyme-activatable ratiometric near-infrared (NIR) fluorescent probes enabling noninvasive imaging of enzyme activity in vivo are promising for biomedical research; however, such probes with ratiometric fluorescence emissions both in NIR window under a single NIR light excitation are largely unexplored. Here, a quenched NIR fluorophore of Cy5.5 is integrated with NIR fluorescent poly[2,6-(4,4-bis-(2-ethylhexyl)-4H-cyclopenta[2,1-b;3,4-b']dithiophene)-alt-4,7(2,1,3-benzothiadiazole)] (PCPDTBT)-based semiconducting polymer nanoparticles (SPNs), and an αv β3 integrin-targeting and matrix metalloproteinase-2 (MMP-2)-activatable ratiometric fluorescent probe (SPN-MMP-RGD) is developed. Under excitation at 660nm, SPN-MMP-RGD shows "always-on" fluorescence of PCPDTBT (830nm) and activatable fluorescence of Cy5.5 (690nm) toward MMP-2, affording a remarkable ≈176-fold enhancement in fluorescence intensity ratio between 690 and 830nm (I690 /I830 ) for sensitive detection of MMP-2 activity in vitro and in tumor cells. By virtue of ratiometric fluorescence imaging independently of probe's concentration, SPN-MMP-RGD can not only accurately report on MMP-2 levels regarding different tumor sizes, but also noninvasively delineate MMP-2-positive tiny gastric tumors metastasis in vivo. The authors' study reveals the potential of SPN-MMP-RGD for ratiometric fluorescence imaging of MMP-2 activity via combining two independent NIR fluorophores, which can be amenable for the design of other enzyme-activatable ratiometric NIR fluorescent probes for reliable in vivo imaging.

Design and Synthesis of New Acridone-Based Nitric Oxide Fluorescent Probe.

Nitric oxide (NO) is an important signaling molecule involved in a wide range of physiological and pathological processes. Fluorescent imaging is a useful tool for monitoring NO concentration, which could be essential in various biological and biochemical studies. Here, we report the design of a novel small-molecule fluorescent probe based on 9(10H)acridone moiety for nitric oxide sensing. 7,8-Diamino-4-carboxy-10-methyl-9(10H)acridone reacts with NO in aqueous media in the presence of O2, yielding a corresponding triazole derivative with fivefold increased fluorescence intensity. The probe was shown to be capable of nitric oxide sensing in living Jurkat cells.

Carbon dots inspired by structure-inherent targeting for nucleic acid imaging and localized photodynamic therapy

Targeting capability is one of the most important properties of both imaging and therapeutic agents. The discovery of “structure-inherent targeting” (SIT) phenomena has benefitted the tailored design of fluorescent probes and photodynamic photosensitizers. Herein, SIT was combined with the inheritance of epigenetic characteristics from parent carbon sources to synthesize nucleic acid-targeted carbon dots (CDs) from quinoline derivatives, namely, 1-CDs, Cl-CDs, and I-CDs via a bottom-up strategy. These CDs exhibited RNA-selective fluorescence imaging, as confirmed using cell digestion, SYTO® RNASelect™ costaining, and nucleic acid titration. Furthermore, owing to iodine doping, I-CDs exhibited excellent PDT performance for killing cancer cells in vitro. Importantly, I-CDs could actively target and become enriched at tumor sites using folic acid-functionalized liposomes as tumor-homing vehicles, thus effectively inhibiting tumor growth in vivo. By screening the SIT carbon source, photodynamic CDs can provide a new platform for next-generation nucleic acid-targeting photodynamic therapy.

Front Cover: Design, Synthesis, and Biological Evaluation of Novel Fluorescent Probes Targeting the 18‐kDa Translocator Protein (12/2021)

The Front Cover illustrates a newly synthesized fluorescent probe for the visualization of 18 kDa translocator protein (TSPO). It exhibits a subnanomolar binding affinity (Ki=0.58 nm) and is partially colocalized with TSPO in brain sections of lipopolysaccharide (LPS)-injected mice. It possesses favorable fluorescence properties, especially in nonpolar environments and could be used to visualize TSPO overexpression in neuroinflammation and degeneration. Cover design by Hendris Wongso. More information can be found in the Full Paper by Hendris Wongso, Andrew Katsifis, Paul A. Keller et al..

Rational design of AIE-based fluorescent probes for hypochlorite detection in real water samples and live cell imaging

As one kind of important disinfectant and reactive oxygen species (ROS), hypochlorite (ClO-), plays vital roles in both water treatment and cell homeostasis. In this work, by decorating a series of groups with different electron donating and withdrawing properties on tetraphenylethene (TPE), four aggregation-induced emission (AIE)-based fluorescent probes containing C˭C double bonds as the potential reaction sites named Probe A, B, C and D were constructed, and their sensing performance for ClO- was systematically studied. The results showed that the substituents can not only effectively tune the photophysical properties of the probes, but also make a significant impact on their sensing performance for ClO-. Combined with the theoretical calculation results, it can be inferred that the reactivity of the probes for ClO- can be greatly enhanced with the increase of electron cloud density on the C˭C double bonds by the introduction of strong electron-donating group (EDG) and electron-withdrawing group (EWG) adjacent to the double bonds. Finally, the best performing Probe D was selected and then successfully applied to ClO- detection in real water samples and live cell imaging.