Mitochondria-targeted Probe Research Articles

Construction of a mitochondria-targeted probe to monitor cysteine levels in cancer cells and zebrafish.

Cysteine (Cys) plays an indispensable role as an antioxidant in the maintenance of bioredox homeostasis. We have constructed an efficient fluorescent probe Mito-Cys based on the binding of indole and naphthol. The acrylic ester group serves as a recognition switch for specific detection of Cys, which undergoes Michael addition and intramolecular cyclization reactions, thereby ensuring the chemical kinetics priority of Cys compared to other biothiols. The probe has good water solubility, large Stokes shift (137nm), with a detection limit of 21.81nM. In addition, cell imaging experiments have shown that the probe has excellent mitochondrial targeting ability (R = 0.902). The probe can distinguish between Cys, homocysteine (Hcy) and glutathione (GSH), and can detect Cys specifically and quickly (100s) to ensure accurate quantitative analysis of Cys changes in cells. More importantly, the probe confirms that ferroptosis inducing factors trigger thiol starvation in mitochondria, which helps to gain a deeper understanding of the physiological and pathological functions related to Cys and ferroptosis.

Rapid and accurate identification of stem cell differentiation stages via SERS and convolutional neural networks

Monitoring the transition of cell states during induced pluripotent stem cell (iPSC) differentiation is crucial for clinical medicine and basic research. However, both identification category and prediction accuracy need further improvement. Here, we propose a method combining surface-enhanced Raman spectroscopy (SERS) with convolutional neural networks (CNN) to precisely identify and distinguish cell states during stem cell differentiation. First, mitochondria-targeted probes were synthesized by combining AuNRs and mitochondrial localization signal (MLS) peptides to obtain effective and stable SERS spectra signals at various stages of cell differentiation. Then, the SERS spectra served as input datasets, and their distinctive features were learned and distinguished by CNN. As a result, rapid and accurate identification of six different cell states, including the embryoid body (EB) stage, was successfully achieved throughout the stem cell differentiation process with an impressive prediction accuracy of 98.5%. Furthermore, the impact of different spectral feature peaks on the identification results was investigated, which provides a valuable reference for selecting appropriate spectral bands to identify cell states. This is also beneficial for shortening the spectral acquisition region to enhance spectral acquisition speed. These results suggest the potential for SERS-CNN models in quality monitoring of stem cells, advancing the practical applications of stem cells.

Engineering a “turn-on” NIR fluorescent sensor-based hydroxyphenyl benzothiazole with a cinnamoyl unit for hydrazine and its environmental and in-vitro applications

Hydrazine (N2H4), a chemical compound widely used in various industrial applications, causes significant environmental and biological hazards. Therefore, it is crucial to develop methodologies for the visualization and real time tracking of N2H4. In this regard, we have constructed a novel near-infrared fluorescent probe (HBT-Cy) that can effectively detect N2H4 in various samples. HBT-Cy contains 2-(2′-hydroxyphenyl)benzothiazole (HBT), cinnamoyl (Cy), and pyridinium (Py) moieties. Importantly, HBT-Cy exhibits a rapid, selective, and highly sensitive response to N2H4. This response results in the release of HBT-Py and the generation of considerable colorimetric changes along with a significant NIR (near infrared) fluorescence signal, peaking at 685 nm. Advantages of this system include turn on NIR fluorescence with large Stokes shift, (approximately 171 nm), low limit of detection (LOD = 0.11 μM) and quantum yield (0.211). The probe with low cytotoxic behavior demonstrates strong NIR fluorescence imaging capabilities to visualize endogenous and exogenous N2H4 in live cells. This mitochondria-targetable probe shows effective subcellular localization. These results suggest that HBT-Cy is a valuable probe for tracking and investigating the behavior of N2H4 in biological systems and environmental samples.

Versatile side chain turns property and functions of α-cyanostilbene derivatives with near-infrared emission and two-photon property for mitochondrial imaging

Developing red materials with bright near-infrared emission and excellent two-photon performance by a simple preparation process is a meaningful and challenging work. Herein, a series of nitro functionalized D-π-A type α-cyanostilbene derivatives with different side chains (ZH, ZOM, ZOE, and ZOB) have been designed and synthesized by a one-pot method. The compounds show weak emissions and almost the same photophysical properties in solutions. In contrast, in the aggregation state, ZOM, ZOE, and ZOB with side chains of methoxy, ethoxy and benzyloxy emit strong fluorescence, while ZH with no side chain shows weak emission. The aggregate emission mechanism is ascribed to the side chain induced the molecular planarization, and formation of unique “bricks” H-aggregates and “staircase” J-aggregates, investigated by single-crystal analysis and DFT calculations. Considering the excellent photophysical properties of the side chain modified compounds, the triphenylphosphonium is employed to the side chain to obtain a mitochondrial targeting probe Z-TPP, which performs high fluorescence and selectively targeted the mitochondria in live cell and zebrafish under one/two-photon excitation, and further successfully applied to observe the complex mitochondrial network with high signal-to-noise ratio under stimulated emission depletion (STED) nanoscopy.

A novel pathway for ratiometric hydrazine sensing in environmental samples and the detection of intracellular viscosity by a mitochondria-targeted fluorescent sensor

Mass and signal transfer, dispersion of reactive metabolites in living cells, and interactions between biomacromolecules are greatly affected by viscosity inside the cells. It is crucial to accurately determine viscosity for reliable results because of the complexities of live cells. Herein, we introduce a new fluorescence probe based on the cyanobiphenyl and benzothiazolium units. This probe not only responds to intracellular viscosity but also detects hydrazine, a widely used chemical that poses significant environmental and toxic risks to organisms. The proposed sensing mechanism provides a new pathway that includes intramolecular cyclization with hydrazine, which differs from other sensing mechanisms. A weak emission (at 590 nm) of the probe under excitation at 365 nm resulted in 25-fold higher emission at 488 nm after the addition of N2H4. The quantum yield of the probe (Φ = 0.089) increased to Φ = 0.199 with the addition of N2H4. In addition, the probe demonstrated 45-fold emission enhancement at 560 nm in viscous media, with a color change from non-fluorescence to yellow fluorescence. Good hydrazine sensing features with high adaptability, selectivity, sensitivity, ratiometric and fast response (90 s), low cytotoxicity (more than 90% of cell viability), low detection limit (86.0 nM), good linearity in the range of 0–35.0 μM, and high signal-to-noise ratio sensing capability were achieved. The hydrazine-sensing capability of the mitochondria-targetable probe in living cells makes it a strong candidate for various biological and environmental applications, including intracellular tracking and imaging. These results suggest that the present probe shows significant potential for the effective fluorescence detection of hydrazine.

Mitochondrial sensitive probe with aggregation‐induced emission characteristics for early brain diagnosis of Parkinson's disease

AbstractThe early diagnosis of Parkinson's disease (PD) provides opportunities for early intervention to slow the progression of neurological degeneration in patients, particularly as the aging population increases in our society. Among a series of pathological features of PD, mitochondria abnormalities have been identified as central event that occurs at the early stage of PD. However, the method for detecting mitochondrial abnormalities‐associated early PD has not been fully developed. We herein report a specifically mitochondrial targeting probe (named TPA‐BT‐SCP) that is able to characterize mitochondria abnormalities for early diagnosis of PD and monitor PD neurodegenerative progress. The probe is an aggregation‐induced emission (AIE) probe with a strong positive charge, a 3D distorted molecular structure, and a separated HOMO‐LUMO distribution, designed with unique molecular design guidelines. Our research demonstrated that TPA‐BT‐SCP could emit stable and strong fluorescence, and rapidly accumulate in mitochondria due to the negative charge. After intranasal administration of 1‐methy‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP)‐induced PD mice, TPA‐BT‐SCP successfully bypassed the blood−brain barrier to light up the brain, allowing the grading of PD severity based on its high sensitivity. Taken together, this work develops a novel AIE probe that exhibits dramatically high sensitivity to mitochondrial changes and enables noninvasive diagnosis of early PD in the brain.

Orange-Emissive Carbon Dots with High Photostability for Mitochondrial Dynamics Tracking in Living Cells.

Mitochondria play significant roles in maintaining a stable internal environment for cell metabolism. Hence, real-time monitoring of the dynamics of mitochondria is essential for further understanding mitochondria-related diseases. Fluorescent probes provide powerful tools for visualizing dynamic processes. However, most mitochondria-targeted probes are derived from organic molecules with poor photostability, making long-term dynamic monitoring challenging. Herein, we design a novel mitochondria-targeted probe based on carbon dots with high performance for long-term tracking. Considering that the targeting ability of CDs is related to surface functional groups, which are generally determined by the reaction precursors, we successfully constructed mitochondria-targeted O-CDs with emission at 565 nm through solvothermal treatment of m-diethylaminophenol. The O-CDs are bright with a high quantum yield of 12.61%, high mitochondria-targeting ability, and good stability. The O-CDs possess a high quantum yield (12.61%), specific mitochondria-targeting ability, and outstanding optical stability. Owing to the abundant hydroxyl and ammonium cations on the surface, O-CDs showed obvious accumulation in mitochondria with a high colocalization coefficient of up to 0.90 and remained steady even after fixation. Besides, O-CDs showed outstanding compatibility and photostability under various interruptions or long-time irradiation. Therefore, O-CDs are preferable for the long-term tracking of dynamic mitochondrial behavior in live cells. We first observed the mitochondrial fission and fusion behaviors in HeLa cells, and then, the size, morphology, and distribution of mitochondria in physiological or pathological conditions were clearly recorded. More importantly, we observed different dynamics interactions between mitochondria and lipid droplets during the apoptosis and mitophagy processes. This study provides a potential tool for exploring interactions between mitochondria and other organelles, further promoting the research on mitochondria-related diseases.

Precise visualization and ROS-dependent photodynamic therapy of colorectal cancer with a novel mitochondrial viscosity photosensitive fluorescent probe

BackgroundColorectal cancer (CRC) is a prominent global cancer with high mortality rates among human beings. Efficient diagnosis and treatment have always been a challenge for CRC management. Fluorescence guided cancer therapy, which combines diagnosis with therapy into one platform, has brought a new chance for achieving precise cancer theranostics. Among this, photosensitizers, applied in photodynamic therapy (PDT), given the integration of real-time imaging capacity and efficacious treatment feasibility, show great potential to serve as remarkable tools. Although much effort has been put into constructing photosensitizers for locating and destroying CRC cells, it is still in high need to develop novel photosensitizers to attain specific detection and fulfil effective therapy.MethodsProbe HTI was rational synthesized for the diagnosis and treatment of CRC. Spectrometric determination was carried out first, followed by the 1O2 generation ability test. Then, HTI was displayed in distinguishing CRC cells from normal cells Further, the PDT effect of the photosensitizer was studied in vitro. Additionally, HTI was used in CRC BALB/c nude mice model to validate its viscosity labelling and tumor suppression characteristics.ResultsWe successfully fabricated a mitochondrial targeting probe, HTI, together with remarkable viscosity sensitivity, ultralow background interference, and excellent 1O2 generation capacity. HTI was favorably applied to the viscosity detection, displaying a 11-fold fluorescent intensity enhancement in solvents from 1.57 cp to 2043 cp. Then, it was demonstrated that HTI could distinguish CRC cells from normal cells upon the difference in mitochondrial viscosity. Moreover, HTI was qualified for producing 1O2 with high efficiency in cells, supported by the sparkling signals of DCFH after incubation with HTI under light irradiation. More importantly, the viscosity labelling and tumor suppression performance in CRC CDX model was determined, enriching the multifunctional validation of HTI in vivo.ConclusionsIn this study, HTI was demonstrated to show a sensitive response to mitochondrial viscosity and possess a high 1O2 generation capacity. Both in vitro cell imaging and in vivo tumor treatment trials proved that HTI was effectively served as a robust scaffold for tumor labeling and CRC cells clearance. This breakthrough discovery held immense potential for advancing the early diagnosis and management of CRC through PDT. By leveraging HTI's properties, medical professionals could benefit from improved diagnostic accuracy and targeted treatment in CRC management, ultimately leading to enhanced patient outcomes.

Meso pyridinium BODIPY-based long wavelength infrared mitochondria-targeting fluorescent probe with high photostability.

Mito-tracker deep red (MTDR) as a commercially available mitochondria-targeting probe was easily bleached upon imaging. We designed and synthesized a family of meso-pyridinium BODIPY and introduced lipophilic methyl or benzyl as the head moiety to develop a mitochondria-targeting deep red probe. Moreover, we changed the substitution of the 3,5-phenyl moieties with the methoxy or methoxyethoxyethyl group to balance hydrophilicity. The designed BODIPY dyes possessed long absorption and good fluorescence emission. Among them, meso ortho-pyridinium BODIPYs with benzyl head and glycol substitution on phenyl moiety (3h) with favorable Stokes shift were found to have the best mitochondrial targeting performance. 3h was easily uptaken by cells and was less toxic and more photostable than MTDR. An immobilizable probe (3i) was further developed, and nice mitochondria targeting properties under the damaging condition of mitochondria membrane potential were maintained. BODIPY 3h or 3i may become alternative long-wavelength mitochondria targeting probes apart from MTDR and be suitable for long-term mitochondrial tracking studies.

Fast responding of mitochondrial HClO using a coumarin-based fluorescent probe in living cells and zebrafish

As an important reactive oxygen species (ROS), hypochlorous acid (HClO) plays pivotal role in the human immune system and ensures the normal function of the organisms. HClO is derived from mitochondria and can regulate intracellular redox balance. Nevertheless, abnormal HClO levels is related to immune system disorders, neurodegenerative injury and cancer. Therefore, fast and sensitive monitoring HClO in mitochondria is extremely significant for some diseases indication. Herein, we designed a mitochondrial-targeted probe HDCl to detect HClO in living cells, which was a hybrid of coumarin, quaternized pyridine (mitochondrial target site) and N, N-dimethylthiocarbamate (DMTC, hypochlorous acid recognition site). The probe HDCl present high sensitivity, high selectivity and ultrafast (<5 s) responding properties for HClO in phosphate buffer solution (PBS) with the very low detection limit of 2.8 nM and trigger a significant enhanced fluorescent signal at 641 nm. Additionally, with a massive Stokes shift (176 nm), HDCl has been successfully applied to imaging HClO in mitochondria of living cells and in zebrafish.

Photocontrollable Fluorescence Imaging of Mitochondrial Peroxynitrite during Ferroptosis with High Fidelity.

Ferroptosis, a new regulatory cell death modality, underlies the pathogenesis of a broad range of disorders. Although much efforts have been made to uncover the molecular mechanisms, some mechanistic details of ferroptosis still remain poorly understood. Particularly, the functional relevance of mitochondrial reactive oxygen species (ROS) in ferroptosis is still highly controversial, which is partially due to the fact that it still remains puzzled how the mitochondrial ROS level varies during ferroptosis. The conventional mitochondria-targeted probes may react with cytosolic ROS and show fluorescence variation before entering mitochondria, thus probably giving a false result on the mitochondrial ROS level and leading to the misjudgment on its biofunction. To circumvent this issue, we rationally designed a photocontrollable and mitochondria-targeted fluorescent probe to in situ visualize the mitochondrial peroxynitrite (ONOO-), which is the ROS member and mediator of ferroptosis. The photoactivated probe was endowed with a highly specific and sensitive fluorescence response to ONOO-. Notably, the response activity could be artificially regulated with light irradiation, which ensured that all the probe molecules passed through the cytosol in the locked status and were then photoactivated after reaching mitochondria. This photocontrolled fluorescence imaging strategy eliminated the interference of ONOO- outside the mitochondria, thus potentially afforded improved fidelity for mitochondrial ONOO- bioimaging in live cells and animal models. With this probe, for the first time, we revealed the mitochondrial ONOO- flux and its probable biological source during erastin-induced ferroptosis. These results suggest a tight correlation between mitochondrial ONOO-/ROS and ferroptotic progression, which will further facilitate the comprehensive exploration and manipulation of ferroptosis.

A near-infrared and lager stocks shift xanthene-indolium sensor for probing hydrazine in mitochondria

Hydrazine (N2H4), which can disrupt the function of mitochondria and damage the morphology of mitochondria, leading to serious diseases, was classified as a restricted substance in drinking water by the U.S. Environmental Protection Agency (EPA) and the World Health Organization (WHO). In this paper, PHNN, a mitochondrial targeting probe with near-infrared emission wavelength and large Stokes shift was constructed for hydrazine detection through the expansion of the π-conjugate system, heteroatom replacement and the introduction of nitrogen positive ions. Surprisingly, the fluorescence emission wavelength of PHNN was extended to 803 nm, and the Stokes shift was widened to 158 nm, respectively. PHNN can specifically recognize hydrazine with high sensitivity with the detection limit lower than the maximum limit of hydrazine in drinking water of EPA and WHO. Importantly, the rationally designed probe PHNN could be used for imaging hydrazine in the mitochondria of living cells, providing a potential tool for further exploration of the toxicity of hydrazine to mitochondria via visualize methods.

PEGylated near-infrared fluorescence probe for mitochondria-targetable hydrogen peroxide detection

Oxidative stress plays significant roles in the development of various diseases. H2O2 acts as a signaling molecule physiologically or harmful substance pathologically and the mitochondria are one of the most active places for the generation of H2O2. Thus, a new mitochondria-targeted probe 1 for H2O2 detection was synthesized herein, based on D-π-A structure with a large Stokes shift (150 nm) due to its ICT process. To improve its water solubility and sensitivity, probe 2 with PEG chain and probe 3 with two responsive boronated groups were then designed based on the structure of probe 1. As a result, the fluorescence intensity of probe 2 was far higher than that of probe 1 and probe 3 not only in vitro experiment but in cell imaging study with a larger linear range and signal-to-noise ratio, rendering it the best probe for further exogenous and endogenous H2O2 detection in Hela cells.

Novel meso-trifluoromethyl BODIPY-based near-infrared-emitting fluorescent probes for organelle-specific imaging of cellular viscosity

Fluorescent molecular rotors play vital roles in understanding the function of cellular viscosity changes and its abnormal fluctuations are closely associated with many diseases. Despite numerous efforts spent, it remains an urgent and important topic to develop improved viscosity probes with near-infrared fluorescence, high sensitivity and low background. In this manuscript, for the first time, we reported two novel monostyryl boron dipyrromethenes (BODIPYs) bearing a rotatable trifluoromethyl group (CF3) at the meso position and a triphenylphosphonium (4) or morpholine (6) substituent, which could successfully serve as highly sensitive and near-infrared-emitting fluorescent probes for imaging cellular viscosity in mitochondria or lysosome. These probes could be prepared through Knoevenagel condensation under mild conditions and showed negligible emissions in the low viscous solvents. However in high viscous glycerol, strong near-infrared fluorescence at 657 nm appeared, ascribed to the substantially restricted rotation of the CF3 group. Their fluorescence intensity (log I) at 657 nm exhibited a good linear relationship with viscosity (log η) in the whole test range of 1–950 cP. Limit of detection for viscosity of mitochondria-targeting probe 4 was relatively lower than that of lysosome-targeting probe 6. These probes both showed no cytotoxicity and good mitochondrial or lysosomal localization. Interestingly, mitochondria-targeting probe 4 exhibited a remarkable intracellular fluorescence enhancement with viscosity increase induced by LPS or nystatin, which was better than lysosome-targeting probe 6. These two novel BODIPY-based fluorescent rotors provide a new strategy for developing near-infrared-emitting and organelle-specific fluorescent probes for cellular viscosity, which might be useful in exploring the functions of viscosity in various diseases.

A mitochondria-targeted probe containing multi-rotors for visualizing the viscosity change in living cells

Fluorescent probes based on molecular rotor are expected to be a powerful tool to uncover the relationship between intracellular viscosity and organelle functions as well as related diseases. Herein, a fluorophore MV-indol containing multiple rotors was constructed by linking triphenylamine group and benzindole unit. It was found that MV-indol can quantitatively detect viscosity with a high selectivity and a 75-fold enhancement in quantum yield (QY). Moreover, the viscosity-sensitive property could be ascribed to the inhibition of molecular rotation and restriction of twisted intramolecular charge transfer (TICT) in a high-viscous environment, which was confirmed by time-dependent density functional theory (TD-DFT). Importantly, we also demonstrated that MV-indol can precisely localize in mitochondria and monitor the change of mitochondrial viscosity at the cellular level.

A ratiometric fluorescence mitochondrial-targeted probe for imaging HOCl in vitro and in vivo

HOCl, as one of reactive oxygen species, could cause tissue damage and induce various disease when it is at abnormal level or improper location. To monitor HOCl, we developed a near infrared ratiometric fluorescence probe C3H, which is capable to detect HOCl in real time. This probe gives two emission signals with no cross-talk in the process of monitoring HOCl, accompanied by a considerable variation (up to 348-fold) in its fluorescence intensity ratio (F480 nm/F750 nm). Additionally, probe C3H exhibited excellent properties such as fast response (<10 s), high sensitivity with a low detection limit (about 22.6 nM), high selectivity and anti-interference. Moreover, the probe possesses the capability of monitoring exogenous and endogenous HOCl in mitochondria. Finally, nude mouse imaging experiments show that the probe can detect exogenous ClO− in vivo.

Mitochondria-targeted acridine-based dual-channel fluorescence chemosensor for detection of Sn4+ and Cr2O72- ions in water and its application in discriminative detection of cancer cells

The goal of the present work was to fabricate a new low-cost, easy-to-prepare, dual-channel fluorescence chemosensor comprised of acridine-diphenylacetyl moieties (NDA) to enable remarkable Sn4+ detection in water and biological medium. The resulting NDA–Sn4+ complex was utilized for the distinguished identification of Cr2O72- ions from other anions and biomolecules. These investigations involve the absorption, fluorescence, and electrochemical methods for the detection of Sn4+ and Cr2O72- ions in pure water. The mechanism for NDA-mediated Sn4+ detection was experimentally determined by FT-IR, NMR titrations, mass (ESI) analyses, and DFT calculations. The obtained results indicate that the NDA chemosensor possessed excellent performance characteristics including good water solubility and compatibility, quick response time (less than 10 s), high sensitivity (Sn4+ = 0.268 μM and Cr2O72- = 0.160 μM), and selectivity against coexisting metals, anions, amino acids, and peptides. The chemosensor NDA induced negligible toxicity in live cells and was successfully utilized as a biomarker for the tracking of Sn4+ in human normal and cancer cells. More importantly, NDA demonstrates distinguished recognition of Sn4+ in human cancer cells rather than in normal live cells. Additionally, NDA was shown to act as a mitochondria-targeted probe in FaDu cells.

Novel Mitochondria-Targeting and Naphthalimide-based Fluorescent Probe for Detecting HClO in Living Cells.

As a key reactive oxygen species (ROS), hypochlorous acid (HClO) plays an important role in many physiological and pathological processes. The mitochondria-targeting probes for the highly sensitive detection of HClO are desirable. In present work, we designed and synthesized an original mitochondria-localizing and turn-on fluorescent probe for detecting HClO. 4-Aminonaphthalimide was employed as the fluorescent section, the (2-aminoethyl)-thiourea unit was utilized as a typical sensing unit, and the quaternized pyridinium moiety was used as a mitochondria-targeted localization group. When HClO was absent, the probe showed weak fluorescence. In the existence of HClO, the probe revealed a blue fluorescence. Moreover, the turn-on fluorescent probe was able to function in a broad pH scope. There was an excellent linearity between the fluorescence emission intensity at 488 nm and the concentrations of HClO in the range of 5.0 × 10–7 to 2.5 × 10–6 mol·L–1. Additionally, the probe had almost no cell toxicity and possessed an excellent mitochondria-localizing capability. Furthermore, the probe was able to image HClO in mitochondria of living PC-12 cells. The above remarkable properties illustrated that the probe was able to determine HClO in mitochondria of living cells.

Revealing Minor pH Changes of Mitochondria by a Highly Sensitive Molecular Fluorescent Probe.

Mitochondrial pH is an important factor associated with cellular metabolism and pathological states. Thus, sensitively monitoring its minor change was essential. However, it was challengeable due to the lack of suitable probes. Here, a mitochondria-targeted probe (NIR-OH-1) was synthesized. Based on the protonation/deprotonation of the hydroxy group and the assistance of carboxyl group on NIR-OH-1 molecular structure, a dramatic NIR activated signal was generated for sensing pH. Probe NIR-OH-1 displayed a good photo-stability and reversibility and could detect pH change without interference by other biologically active species. Importantly, NIR-OH-1 had an appropriate pKa value (7.77) and tiny acid-base transition range, which was allowed to map the small pH changes of cellular mitochondrial. Moreover, NIR-OH-1 was also successfully applied in real-time monitoring mitochondrial pH-related pathological events in living cells under different stimulation, demonstrating the prospect of its clinical application in accurate mitochondrial pH detection under related physiological and pathological conditions.

A mitochondria-targeted fluorescent probe for monitoring endogenous cysteine in living cells and zebrafish.

At the organelle level, pathogenesis due to abnormal concentrations of cysteine (Cys) is of great significance for the early diagnosis and treatment of related diseases. Generally speaking, organelle localization requires the participation of specific target groups, which increases the difficulty of synthesis. Herein, through simple synthesis, a novel biflavone derivative (BFD) that exhibits excited-state intramolecular proton transfer (ESIPT) was obtained and successfully located in mitochondria without target groups. The probe BFD can distinguish Cys from Hcy and GSH with a rapid response (< 5 s) and showed visual detection for Cys with a large Stokes shift (about 260 nm). Because of its nanomorphology in solution and surface functional groups, the probe BFD can enter the cell smoothly and achieve mitochondrial localization. Owing to its excellent optical performance, the probe BFD was successfully applied to the imaging of endogenous Cys in HeLa cells and zebrafish.