Selective Fluorescent Probe Research Articles

Multi-stimuli responsive fluorescence of amphiphilic AIEgen copolymers for ultrafast, highly sensitive and selective copper ion detection in water

Novel multi-stimuli responsive fluorescence sensors based on amphiphilic copolymers containing hydrophilic N-isopropylmethacrylamide (NIPAM) and AIEgenic tetraphenylethylene-dipicolylamine (TPEDPA) monomers attached to side-chains of copolymers are synthesized. Two copolymers poly(NIPAM)x-co-(TPEDPA)y prepared by free radical polymerization with different molar ratios of monomers NIPAM and TPEDPA (i.e., x:y = 60:1 and 175:1 for P1 and P2, respectively) show good solubilities in organic solvents and water, which exhibit strong cyan fluorescence at 466 nm in aggregation and solid states due to AIE behaviors of TPE units. Upon various external stimuli (temperatures and acid/base conditions), P1 and P2 possess excellent reversible thermo- and pH-responsive fluorescence turn-off/on cyan emissions of TPE units. In addition, with DPA receptor, P1 and P2 are applied as ultrafast, selective and sensitive fluorescence probes for copper ion detection in water based on the TPE fluorescence quenching via photoinduced electron transfer (PET) phenomena, which can be recovered by the addition of disodium ethylenediaminetetraacetate solution. Owing to good water solubilities, amphiphilic copolymers P1 and P2 (containing only 1.6 and 0.6 % molar ratios of AIEgen) exhibit significant sensing towards Cu2+ with very prominent limit of detection values (LOD = 57 and 72 nM, respectively), which are much better than that of hydrophobic monomer TPEDPA (590 nM). Moreover, the fluorescence quenching of TPE units due to PET phenomena is further confirmed by theoretical calculations. Additionally, as an excellent quenching fluorescence sensor with good biocompatibility of P1, its recognition of copper ion and bio-imaging applications in living cells are also reported in this research.

Micro flow photochemical synthesis of Ca-sensitive fluorescent sensor particles.

Fluorescence probes have widely been used for detecting and imaging Ca2+‐enriched parts of cells but more rarely for quantitative determination of concentrations. In this study we show how this can be achieved by a novel approach using hydrogel particles. In a microfluidic co‐flow arrangement spherical droplets were generated from an aqueous solution of acrylamide, N,N'‐methylenebisacrylamide crosslinker and photoinitiator and subsequently photo‐cured in situ yielding gel particles in a sub millimeter range. These particles were separated, dried under reduced pressure and re‐swollen in water containing Rhod‐5N tri potassium salt as calcium ion selective fluorescence probe. After that the particles were dried again and stored for further investigations. Upon exposure of dried particles to calcium chloride solutions they swell and take up Ca2+‐ions forming a strong fluorescing complex with Rhod‐5N. Thus, fluorescence intensity increases with calcium ion concentration. Up to ca. 0.50 mM the enhancement effect is strong and then becomes considerably weaker. The intensity‐concentration‐dependence is well described by an equation derived from the equilibrium of the formation of a 1:1 Ca2+:Rhod‐5N complex. The particles allow for a fast optical determination of Ca2+‐concentrations up to 0.50 mM in analyte volumes down to below 10 μL.

Characterization of a Hg2+-Selective Fluorescent Probe Based on Rhodamine B and Its Imaging in Living Cells.

A small organic molecule P was synthesized and characterized as a fluorometric and colorimetric dual-modal probe for Hg2+. The sensing characteristics of the proposed probe for Hg2+ were studied in detail. A fluorescent enhancing property at 583 nm (>30 fold) accompanied with a visible colorimetric change, from colorless to pink, was observed with the addition of Hg2+ to P in an ethanol-water solution (8:2, v/v, 20 mM HEPES, pH 7.0), which would be helpful to fabricate Hg2+-selective probes with “naked-eye” and fluorescent detection. Meanwhile, cellular experimental results demonstrated its low cytotoxicity and good biocompatibility, and the application of P for imaging of Hg2+ in living cells was satisfactory.

A Selective Ratiometric Fluorescent Probe for No-Wash Detection of PVC Microplastic.

Microplastics (MP) are micrometric plastic particles present in drinking water, food and the environment that constitute an emerging pollutant and pose a menace to human health. Novel methods for the fast detection of these new contaminants are needed. Fluorescence-based detection exploits the use of specific probes to label the MP particles. This method can be environmentally friendly, low-cost, easily scalable but also very sensitive and specific. Here, we present the synthesis and application of a new probe based on perylene-diimide (PDI), which can be prepared in a few minutes by a one-pot reaction using a conventional microwave oven and can be used for the direct detection of MP in water without any further treatment of the sample. The green fluorescence is strongly quenched in water at neutral pH because of the formation dimers. The ability of the probe to label MP was tested for polyvinyl chloride (PVC), polyethylene (PE), polyethylene terephthalate (PET), polypropylene (PP), polystyrene (PS), poly methyl methacrylate (PMMA) and polytetrafluoroethylene (PTFE). The probe showed considerable selectivity to PVC MP, which presented an intense red emission after staining. Interestingly, the fluorescence of the MP after labeling could be detected, under excitation with a blue diode, with a conventional CMOS color camera. Good selectivity was achieved analyzing the red to green fluorescence intensity ratio. UV–Vis absorption, steady-state and time-resolved fluorescence spectroscopy, fluorescence anisotropy, fluorescence wide-field and confocal laser scanning microscopy allowed elucidating the mechanism of the staining in detail.

Selective Cadmium Fluorescence Probe Based on Bis-heterocyclic Molecule and its Imaging in Cells.

Fluorescence probes that selectively image cadmium are useful for detecting and tracking the amount of Cd2+ in cells and tissues. In this study, we designed and synthesized a novel Cd2+ fluorescence probe based on the pyridine-pyrimidine structure, 4-(methylsulfanyl)-6-(pyridin-2-yl)pyrimidin-2-amine (3), as a low-molecular-weight fluorescence probe for Cd2+. Compound 3 could successfully discriminate between Cd2+ and Zn2+ and exhibited a highly selective turn-on response toward Cd2+ over biologically related metal ions. The dissociation constant (Kd) and the limit of detection (LOD) of 5.4 × 10- 6mol L- 1 and 4.4 × 10- 7mol L- 1, respectively, were calculated using fluorescence titration experiments. Studies with closely related analogs showed that the bis-heterocyclic moiety of 3 acted as both a coordination site for Cd2+ and a fluorophore. Further, the methylsulfanyl group of compound 3 is essential for achieving selective and sensitive Cd2+ detection. Fluorescence microscopy studies using living cells revealed that the cell membrane permeability of compound 3 is sufficient to detect intracellular Cd2+. These results indicate that novel bis-heterocyclic molecule 3 has considerable potential as a fluorescence probe for Cd2+ in biological applications.

Synthesis of high luminescent Eu3+ doped nanoparticle and its application as highly sensitive and selective detection of Fe3+ in real water and human blood serum

The present work reports a highly efficient Ca doped Eu: Y2O3 i.e Ca0.05Eu0.01Y1.94O3 (CEY.) nanophosphor material synthesized through a facile combustion method, as a simple and selective turn-off fluorescence probe for the quantitative analysis of iron ions (Fe3+). The proposed sensor allows the quantification of iron in the range of 10 µM–90 µM with a limit of detection (LOD) ∼ 63.2 nM under the natural pH range. Moreover, CEY nanophosphor shows an excellent fluorescence phenomenon with a gradual increase in the Fe3+ ion concentration. It has been observed that the corresponding PL intensity gets completely quenched with 500 µM Fe3+ ion concentration. Furthermore, the applicability of the sensor as an efficient probe has been investigated with real water samples, iron tablets, and human blood serum (HBS). The selectivity of the probe has also been analyzed with various metal ions and biomolecules. Thus, in turn, the as-obtained sensing probe illustrates an excellent accuracy, sensitivity, and selectivity, and offers potential application in clinical diagnosis, biological and real water sample studies, with the detection of Fe3+ ion. Furthermore, it does not require any acidic medium for a level-free, and non-enzymic detection of a real sample with almost not affecting the sample quality and henceforth provides more reliable results.

Synthesis and Characterization of a Novel Heterocyclic Schiff Base and Development of a Fluorescent Sensor for Vitamin B12.

A heterocyclic Schiff base (MPDPI)was synthesized by the condensation reaction of 1-phenylisatin with 4,5-dimethylphenylene diamine. It was characterized by using spectroscopic methods including UV visible, Infrared, 1H-NMR, 13C-NMR and mass spectrometry. It acts as the fluorescent probe for the detection of Vitamin B12 (Vit.B12) which shows high selectivity over other species via dynamic quenching mechanism. It is also highly sensitive towards Vit.B12 with a detection limit of [Formula: see text]M and showed a linear concentration ranging from [Formula: see text]to[Formula: see text]. Effect of other coexisting species was also studied. The satisfactory results were also obtained in real samples.Since, there are only few reports on Vit.B12, development of selective fluorescent probes for Vit.B12 would be worthwhile.

In vivo imaging via a red-emitting fluorescent probe to diagnosing liver cancer or drug-induced liver disease

Leucine aminopeptidase (LAP) as an important proteolytic enzyme, has been mainly found in hepatobiliary cells, and overexpressed in hepatoma cells. Herein, a new highly selective red-emitting fluorescent probe (DCDHF-Ala) for LAP has been synthesized based on 2-dicyanomethyldiene-3-cyano-2, 5-dihydrofuran (DCDHF) as fluorophore, and alanine (Ala) as the detection group. More importantly, it's the first time to use Ala as a reactive group for LAP. DCDHF-Ala has a low detection limit (0.20 U/L), excellent water solubility and cell membrane permeability. In addition, the probe has been successfully applied to fluorescent imaging in cells and zebrafish. It's especially worth mentioning that, DCDHF-Ala has a high biosafety and enables a real-time detection of LAP levels in mice model. What's the most important is that DCDHF-Ala may be an effective tool to qualitatively monitor the upregulation of LAP induced by liver injury and liver cancer.

Discovery of Highly Specific Catalytic-Site-Targeting Fluorescent Probes for Detecting Lysosomal PDE10A in Living Cells.

A challenge for sensors targeting specific enzymes of interest in their native environment for direct imaging is that they rationally exploit a highly selective fluorescent probe with a high binding affinity to provide real-time detection. Immunohistochemical staining, proteomic analysis, or recent enzymatic fluorescent probes are not optimal for tracking specific enzymes directly in living cells. Herein, we introduce the concept of designing a highly effective fluorescent probe (BVQ1814) targeting phosphodiesterase 10A with a highly potent affinity and a >1000-fold subfamily selectivity by gaining insights into the three-dimensional structural information of the active site of the catalytic pocket. BVQ1814 showed an outstanding binding affinity for PDE10A in vitro and specifically detected PDE10A in living cells, indicating that most PDE10A was probably distributed in the lysosomes. We validated the PDE10A distribution in stable mCherry-PDE10A-overexpressing HepG2 cells. This probe delineated the profile of PDE10A in tissue sections and exhibited a remarkable therapeutic effect as a PDE10A inhibitor for treating pulmonary arterial hypertension. This concept will open up a new avenue for designing a highly effective fluorescent probe for tracking receptor proteins by taking full advantage of the structural information in the ligand-binding pocket of the target of interest.

Designing a sustainable fluorescent targeting probe for superselective nucleus imaging

Specific targeting of cell organelles is indispensable for bioimaging and diagnostics but remains a challenge due to the limitation of highly selective fluorescent targeting probes. Herein, we successfully designed a molecular fusion route to controllably synthesize bright orange fluorescent graphene quantum dots (GQDs). The relative nitrogen doping level of the GQDs reached 18.88 at%. The GQDs exhibited long-wavelength excitation fluorescence and photoluminescence (PL) stability. Femtosecond transient absorption spectroscopy elucidated that the sp2 cluster and the surface state's synergistic effect contributed to the PL. Furthermore, the most alluring discovery was that the GQDs had good biocompatibility and an unprecedented targeting capability for superselective nucleus imaging without staining cell organelles. Molecular dynamics simulations revealed that once the GQDs were transported across the plasma membrane, the higher the N-doping ratio of the GQDs was, and the easier the GQDs penetrated the membrane. The reason may stem from the compatibility of –NH2 with lipid molecules, which is higher than that of –OH. Enriched nitrogen doping in GQDs is beneficial for crossing the cell membrane to the target nucleus. Our findings could provide scientific theory and a technical basis to design nucleus-targeting probes in the future.

Highly selective colorimetric fluorescent probe for detecting nitrite in aqueous solution

Nitrite (NO2− ions) is a common pollutant in water and soil environment. Developing a rapid and accurate detection method for NO2− ions is always an important task. Herein, a switch-off colorimetric fluorescent probe P-NO2− was presented as the specific monitoring of NO2− ions in aqueous media. Once NO2− ions was added to the probe P-NO2− solution, the solution color changed very quickly, which was from yellow to colorless. And, the fluorescent intensity (580 nm) decreased obviously. The probe P-NO2− could test NO2− ions quantitatively (0.5–10 μM), and the detection limit was 75 nM according to 3σ/slope. Moreover, this new probe could also be used to determinate NO2− ions in the soil solution samples, river and lake water.

A novel carbazole-based highly sensitive and selective turn-on fluorescent probe for mercury (II) ions in aqueous THF

Based on the known deprotection reaction of thioacetal initiated by Hg2+, a new type of fluorescent probe 3-(4-(1,3-dithiolan-2-yl) phenyl)-9H-carbazole (DTCB) is designed and synthesized successfully. The probe shows a good fluorescence response to Hg2+ in THF-H2O (fw = 5%) mixed system. The addition of Hg2+ greatly enhances the fluorescence intensity of the probe and changed from dark blue to green fluorescence. Within a large Hg2+ concentration range (0–6×10−5 mol·L−1), the fluorescence intensity at 480nm presents good linear with the Hg2+ concentration (R2 = 0.993), and the detection limit of the probe to Hg2+ is below 2.05×10−7 mol·L−1. The DTCB probe shows good selectivity and anti-interference performance, and many metal ions and anions hardly effect on its detection performance. The sensing mechanism is confirmed by theory calculation based on DFT and TD-DFT theories, and the intramolecular charge transfer (ICT) is responsible for the significant fluorescence change of probe DTCB. Meanwhile, the probe is applied to the detection of Hg2+ in real water samples; and it is interesting that the DTCB-based test strip firstly reveals the strong fluorescence from solid-state 3-(4-formylphenyl)-9H-carbazole (FCB) with Hg2+ adding. The results show that the probe based on DTCB has potential application prospect in actual sample analysis.

Berberrubine Phosphate: A Selective Fluorescent Probe for Quadruplex DNA.

A phosphate-substituted, zwitterionic berberine derivative was synthesized and its binding properties with duplex DNA and G4-DNA were studied using photometric, fluorimetric and polarimetric titrations and thermal DNA denaturation experiments. The ligand binds with high affinity toward both DNA forms (Kb = 2–7 × 105 M−1) and induces a slight stabilization of G4-DNA toward thermally induced unfolding, mostly pronounced for the telomeric quadruplex 22AG. The ligand likely binds by aggregation and intercalation with ct DNA and by terminal stacking with G4-DNA. Thus, this compound represents one of the rare examples of phosphate-substituted DNA binders. In an aqueous solution, the title compound has a very weak fluorescence intensity (Φfl < 0.01) that increases significantly upon binding to G4-DNA (Φfl = 0.01). In contrast, the association with duplex DNA was not accompanied by such a strong fluorescence light-up effect (Φfl < 0.01). These different fluorimetric responses upon binding to particular DNA forms are proposed to be caused by the different binding modes and may be used for the selective fluorimetric detection of G4-DNA.

A Sensitive and Selective Fluorescent Probe for Real-Time Detection and Imaging of Hypochlorous Acid in Living Cells.

Hypochlorous acid (HClO), a reactive oxygen species, plays an essential role in the processes of physiology and pathology via reacting with most biological molecules. The abnormal level of HClO may cause inflammation, especially arthritis. To further understand its key role in inflammation, in situ detection of HClO is necessary. Herein, a water-soluble small molecule fluorescent probe (HDI-HClO) is employed to monitor and identify trace amounts of HClO in the biological system. In the presence of HClO, the probe releases a hydroxyl group emitting strong fluorescence because of the restoration of the intramolecular charge transfer process. Furthermore, this probe displays a 150-fold fluorescence enhancement accompanied by a large Stokes shift and a lower detection limit (8.3 nM). Moreover, the probe can make a rapid response to HClO within 8 s, which provides the possibility of real-time monitoring of intracellular HClO. Based on the advantages of rapid dynamics, good water solubility, and excellent biocompatibility, this probe could effectively monitor the fluctuations of exogenous and endogenous HClO in living cells. The fluorescence imaging of HDI-HClO indicated that it is an excellent potential approach for comprehending the relationship between inflammation and HClO.

Synthesis and Characterization of a Mg2+-Selective Probe Based on Benzoyl Hydrazine Derivative and Its Application in Cell Imaging.

A simple benzoyl hydrazine derivative P was successfully synthesized and characterized as Mg2+-selective fluorescent probe. The binding of P with Mg2+ caused an obvious fluorescence enhancement at 482 nm. The fluorescent, UV-vis spectra, 1H-NMR, and IR spectra confirmed the formation of P-Mg2+ complex, and the formation of a 1:1 stoichiometry complex was proved by Job’s plot and mass spectrometry. The recognition mechanism of P to Mg2+ was owing to the photoinduced electron transfer effect (PET). The fluorescent response was linear in the range of 0.9–4.0 µM with the detection limit of 0.3 µM Mg2+ in water–ethanol solution (1:9, v:v, pH10.0, 20 mM HEPES). In addition, the results of cell imaging of Mg2+ in Hl-7701 cells was satisfying.

A Novel Fluorescent 1,10‐Phenanthroline‐2,9‐dicarboxaldehyde‐ 2,5‐diaminoterephthalicacid‐Schiff Base Polymer for Cu2+ Detection

AbstractA novel fluorescent Schiff base polymer (PTDA‐DATA‐SBP) is synthesized by amine aldehyde condensation reaction between 1,10‐phenanthroline‐2,9‐dicarboxaldehyde (PTDA) and 2,5‐diaminoterephthalicacid (DATA) as a selective fluorescent probe for Cu2+ detection. The as‐prepared PTDA‐DATA‐SBP displayed irregular nanospheres with a diameter of about 5 μm. Under the excitation of 338 nm, the PTDA‐DATA‐SBP had a characteristic emission peak at 565 nm. The PTDA groups in PTDA‐DATA‐SBP could coordinate with Cu2+ under help of carboxyl groups of DATA. Owing to the photoinduced electron transfer mechanism, the fluorescence of PTDA‐DATA‐SBP was weakened after the addition of Cu2+ in the presence of common ions. Hence, the PTDA‐DATA‐SBP was exploited as a fluorescent nanosensor for sensing Cu2+. The PTDA‐DATA‐SBP showed high selectivity and fast response towards Cu2+ even in presence of other relevant metal ions. The PTDA‐DATA‐SBP fluorescent sensor has a good linear range of 3.45 nM–8.00 μM, and the limit of detection was as low as 1.15 nM. The proposed PTDA‐DATA‐SBP sensor could also detect Cu2+ in actual samples. The work not only provided a good fluorescent probe for Cu2+ detection but also provided a feasible strategy to construct highly selective fluorescent probes.

Carboxylesterases and arylacetamide deacetylase comparison in human A549, H460, and H727 pulmonary cells

AimsHuman carboxylesterases (CESs) and arylacetamide deacetylase (AADAC) are serine-esterase enzymes catalyzing the hydrolysis of many compounds containing esters, amides, thioesters, or acetyl groups. This study aimed to investigate the presence, kinetic parameters, and inhibition of CES1, CES2, and AADAC in A549, H460, and H727 pulmonary cells in both living cells and S9 fractions. Materials and methodsThe p-nitrophenyl acetate (pNPA) and 4-methylumbelliferyl acetate (4-MUA) were used as non-selective esterase substrates, whereas phenacetin as selective AADAC substrate. CESs activities were also investigated in living cells by cellular bioimaging using selective fluorescent probes. Key findingsAADAC gene was detected in A549 and H460 cells; nevertheless, arylesterase activity was not found in relative S9 fractions. Besides, CES1 and CES2 were expressed to a different extent by all lung cells, and enzymatic activities were quite overlapping each other. All enzymes exhibited a typical Michaelis-Menten saturation curve and, regarding 4-MUA, similar Km values were found in both living cells and S9 fractions. Conversely, kinetic parameters relative to the pNPA hydrolysis by S9 fractions were significantly lower than those detected in living cells. Inhibition studies revealed that 4-MUA hydrolysis was inhibited by bis-p-nitrophenyl phosphate and phenylmethanesulfonyl fluoride more than loperamide; on the contrary, pNPA hydrolysis inhibition was limited with similar inhibition profiles being obtained in both living cells and S9 fractions. The presence of carboxylesterases was definitely confirmed by cellular bioimaging. SignificanceThese findings add information to esterase knowledge in pulmonary cells that could be used as in vitro models for toxicological and pharmacological studies.

A highly selective ratiometric fluorescent probe for doxycycline based on the sensitization effect of bovine serum albumin

Fluorescent probes with in-situ visual feature have received numerous attentions for detecting doxycycline (DC), a semisynthetic tetracycline antibiotic widely used in animal husbandry. However, reported fluorescent probes commonly fail to selectively detect DC among tetracycline antibiotics due to their structural similarity. In this work, bovine serum albumin-capped gold nanoclusters (BSA-AuNCs) were ingeniously used as the ratiometric fluorescent probe for detecting DC over other tetracycline antibiotics through the selective sensitization effect of BSA on DC. After adding DC, the red fluorescence of BSA-AuNCs almost remained unchanged, while the green fluorescence of DC also emerged under the sensitization of BSA. BSA-AuNCs showed the highest response toward DC among tetracycline antibiotics ascribed to the strongest sensitization effect of BSA on DC. BSA-AuNCs also displayed the features of simple synthesis, short response time (1 min) and low detection limit (36 nM). BSA-AuNCs were finally applied to detecting DC in fish samples, and further fabricated into test strips for ease of carrying. Thus, this work proposes an efficient strategy to design fluorescent probe for selectively detecting DC among tetracycline antibiotics.

Highly selective fluorescence probe with peptide backbone for imaging mercury ions in living cells based on aggregation-induced emission effect

Mercury is an anthropogenic toxic heavy metal found in the environment. It is highly desirable to develop a fluorescence probe that can selectively and sensitively detect mercury ions using a turn-on response. This paper reports the successful development of a peptide fluorescence probe, TP-2 (TPE-Trp-Pro-Gln-His-Glu-NH2), which uses aggregation-induced emission effects and high selectivity to detect Hg2+. After fluorescence was activated, Hg2+ was efficiently detected using the change in fluorescence intensity. The detection limit for Hg2+ in the buffer solution was 41 nM (R2 = 0.9952). Owing to its high sensitivity, high cell permeability, and low biotoxicity, the probe could perform live cell imaging under biological conditions. This study demonstrated that TP-2 can detect Hg2+ in complex biological environments.

Fluorescent probes for the detection and imaging of Cytochrome P450

Cytochrome P450 (CYP) enzymes are haem-containing proteins which are mainly involved in the oxidative metabolism of various chemicals. CYP enzymes can be affected by co-administrated drugs, environmental factors, disease states and genetic polymorphism, leading to differing degrees of individual variability of CYP activity which can induce the adverse effect of drugs or result in the failure of a drug therapy with varying severity. In this arena, fluorescent probes can serve as useful molecular tools to selectively detect CYP activity, facilitated by their intrinsic sensitivity and capacity for rapid, real-time and in situ monitoring as well as high-throughput screening. Therefore, the design of enzymatic activity reporters serving as the CYP subtype-specific probes, has stimulated widespread interest. This review covers the design principles for probes and systemically summarizes recent advances in the development of fluorescent probes for CYP isoforms including CYP1A, CYP1A1, CYP2C9, CYP2D6, CYP2J2, CYP3A, CYP3A4, CYP3A7, CYP4A11, CYP4F12 and CYP4F3B, and also probes for detecting the reduction ability of CYP, then categorizes these molecular tools according to the detection strategy or interaction models for the key CYP isoforms, such as dealkylation, hydroxylation or reduction reactions. In addition, challenges and future perspectives in the development of selective and sensitive fluorescent probes of CYP are discussed.