An ICT-based strategy to a colorimetric and ratiometric fluorescence probe for hydrogen sulfide in living cells

We present a colorimetric and ratiometric fluorescent probe Cy-N(3) that exhibits a selective response to H(2)S. The probe employs a near-infrared cyanine as a fluorophore, and is equipped with an operating azide unit. It is readily employed for assessing intracellular H(2)S levels, and confocal ratiometric imaging is achieved successfully.     

A FRET-based approach to ratiometric fluorescence detection of hydrogen peroxide

We report the synthesis, properties, and biological applications of Ratio-Peroxyfluor-1 (RPF1), a new ratiometric fluorescent reporter for hydrogen peroxide. RPF1 is comprised of a two-fluorophore cassette, where the spectral overlap between coumarin donor and fluoran/fluorescein acceptor partners can be controlled by the chemoselective peroxide-mediated deprotection of boronic ester pendants on the acceptor dye. RPF1 features good selectivity for hydrogen peroxide over a variety of reactive oxygen species, including superoxide and nitric oxide, a ca. 8-fold increase in fluorescence intensity ratio (lambda517/lambda464) upon H2O2 reaction, and excitation and emission profiles in the visible region. Experiments with viable yeast mitochondria show that RPF1 can monitor and quantify endogenous production of H2O2, establishing the potential utility of this approach for probing peroxide biology in living systems.     

A dual-fluorophore sensor approach for ratiometric fluorescence imaging of potassium in living cells

Potassium is the most abundant intracellular metal in the body, playing vital roles in regulating intracellular fluid volume, nutrient transport, and cell-to-cell communication through nerve and muscle contraction. On the other hand, aberrant alterations in K⁺ homeostasis contribute to a diverse array of diseases spanning cardiovascular and neurological disorders to diabetes to kidney disease to cancer. There is an unmet need for studies of K⁺ physiology and pathology owing to the large differences in intracellular versus extracellular K⁺ concentrations ([K⁺]_intra = 150 mM, [K⁺]_extra = 3–5 mM). With a relative dearth of methods to reliably measure dynamic changes in intracellular K⁺ in biological specimens that meet the dual challenges of low affinity and high selectivity for K⁺, particularly over Na⁺, currently available fluorescent K⁺ sensors are largely optimized with high-affinity receptors that are more amenable for extracellular K⁺ detection. We report the design, synthesis, and biological evaluation of Ratiometric Potassium Sensor 1 (RPS-1), a dual-fluorophore sensor that enables ratiometric fluorescence imaging of intracellular potassium in living systems. RPS-1 links a potassium-responsive fluorescent sensor fragment (PS525) with a low-affinity, high-selectivity crown ether receptor for K⁺ to a potassium-insensitive reference fluorophore (Coumarin 343) as an internal calibration standard through ester bonds. Upon intracellular delivery, esterase-directed cleavage splits these two dyes into separate fragments to enable ratiometric detection of K⁺. RPS-1 responds to K⁺ in aqueous buffer with high selectivity over competing metal ions and is sensitive to potassium ions at steady-state intracellular levels and can respond to decreases or increases from that basal set point. Moreover, RPS-1 was applied for comparative screening of K⁺ pools across a panel of different cancer cell lines, revealing elevations in basal intracellular K⁺ in metastatic breast cancer cell lines vs. normal breast cells. This work provides a unique chemical tool for the study of intracellular potassium dynamics and a starting point for the design of other ratiometric fluorescent sensors based on two-fluorophore approaches that do not rely on FRET or related energy transfer designs.

We report a dual-fluorophore approach for ratiometric fluorescent imaging of K⁺ levels in live cells. Intracellular esterases cleave RPS-1 to detach the K⁺-responsive fluorophore (PS525) from its internal standard (Coumarin 343).     

Development of an ICT-based ratiometric fluorescent hypochlorite probe suitable for living cell imaging

We have judiciously constructed a novel ICT-based ratiometric OCl(-) probe capable of ratiometric imaging in the live cells based on the new OCl(-)-promoted de-diaminomaleonitrile reaction.     

Sulfonate-based fluorescent probes for imaging hydrogen peroxide in living cells

Based on the mechanism of H₂O₂-mediated hydrolysis of sulfonates, two fluorescein disulfonates compounds (FS-1 and FS-2) were designed and synthesized as the highly selective and sensitive fluorescent probes for imaging H₂O₂ in living cells. The probes were detected with elemental analysis, IR, ¹H NMR and ¹³C NMR. Upon reaction with H₂O₂, the probes exhibit strong fluorescence responses and high selectivity for H₂O₂ over other reactive oxygen species and some biological compounds. Furthermore, the sulfonate-based probes, as novel fluorescent reagents, are cell-permeable and can detect micromolar changes in H₂O₂ concentrations in living cells by using confocal microscopy. Supported by the National Basic Research Program of China (Grant No. 2007CB936000), the National Natural Science Funds for Distinguished Young Scholar (Grant No. 20725518), Major Program of the National Natural Science Foundation of China (Grant No. 90713019), the National Natural Science Foundation of China (Grant No. 20875057), the Natural Science Foundation of Shandong Province, China (Grant No. Y2007B02), and the Science and Technology Development Programs of Shandong Province, China (Grant No. 2008GG30003012)     

基于磺酸酯的荧光探针用于活细胞内过氧化氢的成像检测

基于过氧化氢（H2O2）特异性催化水解磺酸酯,设计合成了新型绿色荧光探针：荧光素二磺酸酯（FS—1）和二氯荧光素二磺酸酯（FS-2）两种螺环内酯型化合物,用于活细胞内过氧化氢的检测．探针结构由元素分析、IR、^1H NMR及^13C NMR表征．实验表明：探针FS-1和FS-2在模拟生物体系中检测过氧化氢具有良好的选择性和灵敏度,且线性范围较宽．细胞成像显示：探针FS-1和FS-2用于PMA刺激或外加不同浓度H2O2孵育的小鼠腹膜巨噬细胞均呈现明亮的绿色荧光,且能对细胞内H2O2微摩尔级浓度变化产生响应,证明两探针均具有良好的膜渗透性、高的选择性及良好的灵敏度．该方法的建立对研究生物体内H2O2的产生,H2O2导致的各种疾病机制及H2O2介导的信号转导途径具有重要的理论及实际意义．     

An ICT-based ratiometric probe for hydrazine and its application in live cells

Hydrazine is an important industrial chemical but also very toxic thus requiring rapid detection agents. A ratiometric fluorescence probe that enables rapid, low-limit and naked-eye detection is successfully designed and used for hydrazine determination in live cells.     

A two-photon fluorescent probe for ratiometric imaging of hydrogen peroxide in live tissue

We report a two-photon fluorescent probe (PN1) that can be excited by 750 nm femto-second pulses, shows high photostability and negligible toxicity, and can visualize H(2)O(2) distribution in live cells and tissue by two-photon microscopy.     

A targetable fluorescent probe for imaging hydrogen peroxide in the mitochondria of living cells

We present the design, synthesis, and biological applications of mitochondria peroxy yellow 1 (MitoPY1), a new type of bifunctional fluorescent probe for imaging hydrogen peroxide levels within the mitochondria of living cells. MitoPY1 combines a chemoselective boronate-based switch and a mitochondrial-targeting phosphonium moiety for detection of hydrogen peroxide localized to cellular mitochondria. Confocal microscopy and flow cytometry experiments in a variety of mammalian cell types show that MitoPY1 can visualize localized changes in mitochondrial hydrogen peroxide concentrations generated by situations of oxidative stress.     

A mitochondria-localized two-photon fluorescent probe for ratiometric imaging of hydrogen peroxide in live tissue

We report a two-photon fluorescent probe (SHP-Mito) which can ratiometrically detect mitochondrial H(2)O(2) in live cells and intact tissues at >100 μm depth through the use of two-photon microscopy.     

A ratiometric fluorescent probe for imaging hydroxyl radicals in living cells

A novel fluorescent probe, the detection mechanism of which is based on the 'on-off' switching of a FRET triggered by the *OH-induced cleavage of a DNA strand, has been developed for the ratiometric imaging of *OH.     

A 4-hydroxynaphthalimide-derived ratiometric fluorescent chemodosimeter for imaging palladium in living cells

A highly selective ratiometric fluorescent chemodosimeter derived from 4-hydroxynaphthalimide was designed and synthesized to image palladium species in living cells by virtue of a palladium-catalyzed depropargylation reaction, and it could monitor three typical palladium species (0, + 2 and + 4) without additional reagents.     

An activatable ratiometric near-infrared fluorescent probe for hydrogen sulfide imaging in vivo

Ratiometric fluorescent probes hold great promise for in vivo imaging; however, stimuli-activatable ratiometric probes with fluorescence emissions in near-infrared(NIR) region are still very few. Herein, we report a hydrogen sulfide(H_2S)-activatable ratiometric NIR fluorescent probe(1-SPN) by integrating a H_2S-responsive NIR fluorescent probe 1 into a H_2S-inert poly[2,6-(4,4-bis-(2-ethylhexyl)-4 H-cyclopenta[2,1-b;3,4-b′]dithiophene)-alt-4,7(2,1,3-benzothiadiazole)](PCPDTBT)-based NIR semiconducting polymer nanoparticle(SPN). 1-SPN shows "always on" PCPDTBT fluorescence at 830 nm and weak probe 1 fluorescence at 725 nm under excitation at 680 nm. The ratio of NIR fluorescence intensities between 725 and 830 nm(I_(725)/I_(830))is small. Upon interaction with H_2S, the fluorescence at 725 nm is rapidly switched on, resulting in a large enhancement of I_(725)/I_(830), which is allowed for sensitive visualization and quantification of H_2S concentrations in living cells. Taking advantage of enhanced tissue penetration depth of NIR fluorescence, 1-SPN is also applied for real-time ratiometric fluorescence imaging of hepatic and tumor H_2S in living mice. This study demonstrates that activatable ratiometric NIR fluorescent probes hold great potential for in vivo imaging.     

A selective, cell-permeable optical probe for hydrogen peroxide in living cells

We present the synthesis, properties, and biological applications of Peroxyfluor-1 (PF1), a new type of optical probe for intracellular imaging of hydrogen peroxide in living biological samples. PF1 utilizes a boronate deprotection mechanism to provide unprecedented selectivity and optical dynamic range for detecting H2O2 in aqueous solution over similar reactive oxygen species including superoxide, nitric oxide, tert-butyl hydroperoxide, and hydroxyl radical. We further demonstrate the value of this reagent for biological applications by imaging changes in [H2O2] in living mammalian cells.     

A highly selective colorimetric and ratiometric fluorescent chemodosimeter for imaging fluoride ions in living cells

A simple but highly selective colorimetric and ratiometric fluorescent chemodosimeter was designed and synthesized to detect fluoride ions (F(-)) in aqueous solution and living cells by virtue of the strong affinity of F(-) toward silicon.     

Chemoselective reduction-based fluorescence probe for detection of hydrogen sulfide in living cells

A selective and sensitive fluorescence probe for hydrogen sulfide (H(2)S) detection was synthesized and evaluated in PBS buffer and fetal bovine serum. The effect of pH on the probe was also studied. In addition, visualization of H(2)S in Hela cells was achieved using confocal laser scanning fluorescence microscopy.     

A fluorescence enhanced probe for sulfur dioxide detection and fluorescence imaging in living cellsEI北大核心CSCD

基于香豆素和苯并吡啶基团,构建了用于二氧化硫(SO_(2))高效检测的荧光探针P1,其化学结构通过核磁氢谱(^(1)H NMR)、碳谱(^(13)C NMR)和高分辨质谱(HR-MS)确证。在缓冲溶液体系中,单独的探针P1具有微弱的荧光,识别SO_(2)后荧光发射强度明显增强,能够实现对SO_(2)的专一性裸眼识别,检出限为126 nmol/L。生物应用实验结果表明,该探针具有较低的细胞毒性,可用于生物活细胞中外源性SO_(2)的荧光成像。     

A rational approach to tuning the pKa values of rhodamines for living cell fluorescence imaging

A novel strategy to systematically tune the pK(a) values of rhodamines is described. This strategy was applied to rationally develop compound 1e with a pK(a) of 6.5, the highest among rhodamine amide derivatives, and it could be employed to detect acidic pH variations in living cells with a turn-on signal.     

Selective turn-on fluorescent probes for imaging hydrogen sulfide in living cells

Hydrogen sulfide (H(2)S) is an important biological messenger but few biologically-compatible methods are available for its detection. Here we report two bright fluorescent probes that are selective for H(2)S over cysteine, glutathione and other reactive sulfur, nitrogen, and oxygen species. Both probes are demonstrated to detect H(2)S in live cells.