Organic Fluorescent Dyes Research Articles

Recent Progress of Hybrid Optical Probes for Neural Membrane Potential Imaging.

The neural membrane potential of nerve cells is the basis of neural activity production, which controls advanced brain activities such as memory, emotion, and learning. In the past decades, optical voltage indicator has emerged as a promising tool to decode neural activities with high-fidelity and excellent spatiotemporal resolution. In particular, the hybrid optical probes can combine the advantageous photophysical properties of different components such as voltage-sensitive molecules, highly fluorescent fluorophores, membrane-targeting tags, and optogenetic materials, thus showing numerous advantages in improving the photoluminescence intensity, voltage sensitivity, photostability, and cell specificity of probes. In this review, the current state-of-the-art hybrid probes are highlighted, that are designed by using fluorescent proteins, organic dyes, and fluorescent nanoprobes as the fluorophores, respectively. Then, the design strategies, voltage-sensing mechanisms and the in vitro and in vivo neural activity imaging applications of the hybrid probes are summarized. Finally, based on the current achievements of voltage imaging studies, the challenges and prospects for design and application of hybrid optical probes in the future are presented.

Flavanone-Based Fluorophores with Aggregation-Induced Emission Enhancement Characteristics for Mitochondria-Imaging and Zebrafish-Imaging.

Fluorophores with aggregation-induced emission enhancement (AIEE) characteristics applied in bioimaging have attracted more and more attention in recent years. In this work, a series of flavanone compounds with AIEE characteristics was developed and applied to fluorescence imaging of mitochondria and zebrafish. The compounds were readily prepared by the thermal dehydration of chalcone that was obtained by the reaction of o-hydroxyacetophenone and benzaldehyde. Two of these compounds showed significant AIEE characteristics by fluorescence performance experiments, including optical spectra, fluorescence spectra, fluorescence quantum yield (φF), fluorescence lifetime, and scanning electron microscopy (SEM). Compared with traditional organic fluorescent dyes, these compounds have high fluorescence emission and high fluorescence quantum yield in solid or aggregated state, which overcomes the shortcoming of aggregation-caused quenching (ACQ). More importantly, the two compounds exhibited low cytotoxicity and good cytocompatibility in A549 lung cells at the experimental concentration range and they specifically targeted mitochondria, which make it of great potential use in mitochondria labeling. In addition, they were embryonic membrane permeable and had different affinities for different tissues and organs of zebrafish, but mainly distributed in the digestive system, providing a basis for the application of such compounds in bioimaging. These AIEE compounds with superior properties could be of great potential use in mitochondria imaging and other in vivo studies.

Microplasma Synthesis of Luminescent N-Doped Graphene Quantum Dots for pH Sensing

Graphene quantum dots (GQDs), the newest type of carbon nanoparticles (CNPs) with size <20 nm, have attracted many research attentions due to the highly tunable photoluminescence (PL) properties, biocompatibility, chemical and photo stability, and low toxicity. Doping GQDs with heteroatoms, such as nitrogen, can increase the PL quantum yield and has strong electron withdrawing ability. Therefore, nitrogen-doped GQDs as fluorescent pH monitor excels in sensitivity, selectivity, rapidity, and applicability comparing with the conventional electrochemical method, especially for bio-related and medical applications. Good biocompatibility and low toxicity also make N-GQDs more preferable to semiconductor quantum dots, organic fluorescent dyes, and noble-metal nanoparticles. As a matter of fact, pH value plays a very important and crucial role in all life forms. A small variations in pH not only endangers the lives of plants and animal but also human beings as it will mainly pollute our drinking water, which may cause some health issues, like physiological dysfunction or diseases. Furthermore, cancer and tumor cells can also be diagnosed through cellular imaging due to the abnormal pH inside human body.Most of the approaches for synthesizing N-GQDs are complex, time-consuming, in need of harsh reaction condition, and costly. Hereby, microplasma emerges as one of the most promising approaches for nanomaterials processing due to its facile, stable, efficient, and relatively low cost. The high electron density provided are very energetic (~10eV), which allows non-thermal dissociation of molecular gases to form high concentrations of reactive radical species, e.g. OH˙, O˙, O2 -˙, and NO˙. These reactive radical species along with electrons are brought into the precursor solutions and will interact with the solution chemical species to form GQDs in a cascaded chemical reactions.Here we report a microplasma synthesis of N-GQDs for pH sensing. N-GQDs with high quantum yield up to 30% have been synthesized via microplasma approach from chitosan. This synthetic method leads to highly crystalline particles with easily tunable properties in high reproducibility from a simple biomass as the sole precursor. Based on protonation and deprotonation of N-GQDs surface functional groups, a PL based pH probe with broad linear range of 1.78-13.56 and high accuracy can be constructed. Moreover, two linear lines relationship between UV absorption and different pH value are also obtained, covering from pH 1.25-10.17 and pH 10.17-13.24. The dual optical sensing characteristics allow for almost covering the entire pH range and better accuracy. Figure 1

Constructing a Local Hydrophobic Cage in Dye-Doped Fluorescent Silica Nanoparticles to Enhance the Photophysical Properties.

Aggregation-caused quenching (ACQ) and poor photostability in aqueous media are two common problems for organic fluorescence dyes which cause a dramatic loss of fluorescence imaging quality and photodynamic therapy (PDT) failure. Herein, a local hydrophobic cage is built up inside near-infrared (NIR) cyanine-anchored fluorescent silica nanoparticles (FSNPs) in which a hydrophobic silane coupling agent (n-octyltriethoxysilane, OTES) is doped into FSNPs for the first time to significantly inhibit the ACQ effect and inward diffusion of water molecules. Therefore, the obtained optimal FSNP-C with OTES-modification can provide hydrophobic repulsive forces to effectively inhibit the π–π stacking interaction of cyanine dyes and simultaneously reduce the formation of strong oxidizing species (•OH and H2O2) in reaction with H2O, resulting in the best photostability (fluorescent intensity remained at 90.1% of the initial value after 300 s of laser scanning) and a high PDT efficiency on two- and three-dimensional (spheroids) HeLa cell culture models. Moreover, through molecular engineering (including increasing covalent anchoring sites and steric hindrance groups of cyanine dyes), FSNP-C exhibits the highest fluorescent intensity both in water solution (12.3-fold improvement compared to free dye) and living cells due to the limitation of molecular motion. Thus, this study provides an effectively strategy by combining a local hydrophobic cage and molecular engineering for NIR FSNPs in long-term bright fluorescence imaging and a stable PDT process.

Dye-Loaded Quatsomes Exhibiting FRET as Nanoprobes for Bioimaging.

Fluorescent organic nanoparticles (FONs) are emerging as an attractive alternative to the well-established fluorescent inorganic nanoparticles or small organic dyes. Their proper design allows one to obtain biocompatible probes with superior brightness and high photostability, although usually affected by low colloidal stability. Herein, we present a type of FONs with outstanding photophysical and physicochemical properties in-line with the stringent requirements for biomedical applications. These FONs are based on quatsome (QS) nanovesicles containing a pair of fluorescent carbocyanine molecules that give rise to Förster resonance energy transfer (FRET). Structural homogeneity, high brightness, photostability, and high FRET efficiency make these FONs a promising class of optical bioprobes. Loaded QSs have been used for in vitro bioimaging, demonstrating the nanovesicle membrane integrity after cell internalization, and the possibility to monitor the intracellular vesicle fate. Taken together, the proposed QSs loaded with a FRET pair constitute a promising platform for bioimaging and theranostics.

Porphyrin-terminated nanoscale fluorescent polyrotaxane as a biodegradable drug carrier for anticancer research

Drug delivery carriers hold tremendous promise for improving cancer treatment, and polyrotaxane (PR) has shown excellent drug-carrying properties. However, there have been some reports that, when used as a drug carrier, water-soluble PR is not easily labeled with organic fluorescent dyes. Herein, we synthesized a drug-loaded fluorescent porphyrin-terminated PR (PR-COOH) which can be used as a tracer material in drug and gene delivery. The structure, morphology and zeta potential of PR-COOH were characterized by nuclear magnetic resonance, high-resolution transmission electron microscopy and zeta potentiometry. In this research, cisplatin (CDDP) is used as a model drug. The zeta potential, drug encapsulation efficiency and drug release of CDDP-loaded PR (PR-COOH-Pt) were studied. Confocal laser scanning microscopy showed that PR-COOH could be internalized by HeLa and CT26 cells. The antitumor efficacy of PR-COOH-Pt was investigated in vitro by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and in vivo by a xenograft tumor model. The results showed that PR-COOH-Pt could significantly inhibit tumor growth; thus PR-COOH-Pt has a promising role in cancer therapy.

Surface passivation of zero-mode waveguide nanostructures: benchmarking protocols and fluorescent labels

Zero mode waveguide (ZMW) nanoapertures efficiently confine the light down to the nanometer scale and overcome the diffraction limit in single molecule fluorescence analysis. However, unwanted adhesion of the fluorescent molecules on the ZMW surface can severely hamper the experiments. Therefore a proper surface passivation is required for ZMWs, but information is currently lacking on both the nature of the adhesion phenomenon and the optimization of the different passivation protocols. Here we monitor the influence of the fluorescent dye (Alexa Fluor 546 and 647, Atto 550 and 647N) on the non-specific adhesion of double stranded DNA molecule. We show that the nonspecific adhesion of DNA double strands onto the ZMW surface is directly mediated by the organic fluorescent dye being used, as Atto 550 and Atto 647N show a pronounced tendency to adhere to the ZMW while the Alexa Fluor 546 and 647 are remarkably free of this effect. Despite the small size of the fluorescent label, the surface charge and hydrophobicity of the dye appear to play a key role in promoting the DNA affinity for the ZMW surface. Next, different surface passivation methods (bovine serum albumin BSA, polyethylene glycol PEG, polyvinylphosphonic acid PVPA) are quantitatively benchmarked by fluorescence correlation spectroscopy to determine the most efficient approaches to prevent the adsorption of Atto 647N labeled DNA. Protocols using PVPA and PEG-silane of 1000 Da molar mass are found to drastically avoid the non-specific adsorption into ZMWs. Optimizing both the choice of the fluorescent dye and the surface passivation protocol are highly significant to expand the use of ZMWs for single molecule fluorescence applications.

Design of AIEgens for near-infrared IIb imaging through structural modulation at molecular and morphological levels

Fluorescence imaging in near-infrared IIb (NIR-IIb, 1500–1700 nm) spectrum holds a great promise for tissue imaging. While few inorganic NIR-IIb fluorescent probes have been reported, their organic counterparts are still rarely developed, possibly due to the shortage of efficient materials with long emission wavelength. Herein, we propose a molecular design philosophy to explore pure organic NIR-IIb fluorophores by manipulation of the effects of twisted intramolecular charge transfer and aggregation-induced emission at the molecular and morphological levels. An organic fluorescent dye emitting up to 1600 nm with a quantum yield of 11.5% in the NIR-II region is developed. NIR-IIb fluorescence imaging of blood vessels and deeply-located intestinal tract of live mice based on organic dyes is achieved with high clarity and enhanced signal-to-background ratio. We hope this study will inspire further development on the evolution of pure organic NIR-IIb dyes for bio-imaging.

AC electrokinetic immobilization of organic dye molecules

The application of inhomogeneous AC electric fields for molecular immobilization is a very fast and simple method that does not require any adaptions to the molecule’s functional groups or charges. Here, the method is applied to a completely new category of molecules: small organic fluorescence dyes, whose dimensions amount to only 1 nm or even less. The presented setup and the electric field parameters used allow immobilization of dye molecules on the whole electrode surface as opposed to pure dielectrophoretic applications, where molecules are attracted only to regions of high electric field gradients, i.e., to the electrode tips and edges. In addition to dielectrophoresis and AC electrokinetic flow, molecular scale interactions and electrophoresis at short time scales are discussed as further mechanisms leading to migration and immobilization of the molecules.Graphical

Hot-Band Anti-Stokes Fluorescence Properties of Alexa Fluor 568

Hot-band absorption and anti-Stokes emission properties of an organic fluorescent dye, Alexa Fluor 568, were characterized and compared with those of Rhodamine 101. The comparison of the properties (e.g., quantum efficiency, spectral distribution, thermal properties, and fluorescence lifetime) between the two dyes confirms that both dyes undergo the same process when excited in the red spectral region. Possible undesirable crosstalk effects and applications in dSTORM microscopy were demonstrated and discussed.

Multicomposite super-resolution microscopy: Enhanced Airyscan resolution with radial fluctuation and sample expansions.

Either modulated illumination or temporal fluctuation analysis can assist super-resolution techniques in overcoming the diffraction limit of conventional optical microscopy. As they are not contradictory to each other, an effective combination of spatial and temporal super-resolution mechanisms would further improve the resolution of fluorescent images. Here, a super-resolution imaging method called fluctuation-enhanced Airyscan technology (FEAST) is proposed, which achieves ~40 nm lateral imaging resolution and is useful for a range of fluorescent proteins and organic dyes. It was demonstrated not only to obtain different subcellular super-resolution images, but also to improve the accuracy of counting the average human epidermal growth factor receptor 2 (HER2) copy number for diagnosis in breast cancer. Furthermore, the combination of FEAST and sample expansion microscopy (Ex-FEAST) improves the lateral resolution to ~26 nm.

Core\u2013shell polymeric nanoparticles co-loaded with photosensitizer and organic dye for photodynamic therapy guided by fluorescence imaging in near and short-wave infrared spectral regions

BackgroundBiodistribution of photosensitizer (PS) in photodynamic therapy (PDT) can be assessed by fluorescence imaging that visualizes the accumulation of PS in malignant tissue prior to PDT. At the same time, excitation of the PS during an assessment of its biodistribution results in premature photobleaching and can cause toxicity to healthy tissues. Combination of PS with a separate fluorescent moiety, which can be excited apart from PS activation, provides a possibility for fluorescence imaging (FI) guided delivery of PS to cancer site, followed by PDT.ResultsIn this work, we report nanoformulations (NFs) of core–shell polymeric nanoparticles (NPs) co-loaded with PS [2-(1-hexyloxyethyl)-2-devinyl pyropheophorbide-a, HPPH] and near infrared fluorescent organic dyes (NIRFDs) that can be excited in the first or second near-infrared windows of tissue optical transparency (NIR-I, ~ 700–950 nm and NIR-II, ~ 1000–1350 nm), where HPPH does not absorb and emit. After addition to nanoparticle suspensions, PS and NIRFDs are entrapped by the nanoparticle shell of co-polymer of N-isopropylacrylamide and acrylamide [poly(NIPAM-co-AA)], while do not bind with the polystyrene (polySt) core alone. Loading of the NIRFD and PS to the NPs shell precludes aggregation of these hydrophobic molecules in water, preventing fluorescence quenching and reduction of singlet oxygen generation. Moreover, shift of the absorption of NIRFD to longer wavelengths was found to strongly reduce an efficiency of the electronic excitation energy transfer between PS and NIRFD, increasing the efficacy of PDT with PS-NIRFD combination. As a result, use of the NFs of PS and NIR-II NIRFD enables fluorescence imaging guided PDT, as it was shown by confocal microscopy and PDT of the cancer cells in vitro. In vivo studies with subcutaneously tumored mice demonstrated a possibility to image biodistribution of tumor targeted NFs both using HPPH fluorescence with conventional imaging camera sensitive in visible and NIR-I ranges (~ 400–750 nm) and imaging camera for short-wave infrared (SWIR) region (~ 1000–1700 nm), which was recently shown to be beneficial for in vivo optical imaging.ConclusionsA combination of PS with fluorescence in visible and NIR-I spectral ranges and, NIR-II fluorescent dye allowed us to obtain PS nanoformulation promising for see-and-treat PDT guided with visible-NIR-SWIR fluorescence imaging.

Chirality Evolution from Sub-1 Nanometer Nanowires to the Macroscopic Helical Structure.

The chirality evolution from the molecular level to the macroscopic level remains elusive for inorganic hierarchical structures. Without adding any chiral ligands or dopants, we prepared the macroscopic helical assemblies of sub-1 nm nanowires through a facile evaporation-induced self-assembly process with 100% efficiency, benefiting from the self-adjustment and self-recognition of sub-1 nm nanowires. Furthermore, we observed circularly polarized luminescence signals from the helical assemblies composed of nanowires and achiral organic fluorescent dyes, stemming from chirality transfer from the helical assemblies to achiral organic molecules. Molecular dynamics simulations found that the chirality of nanowires played a key role in the formation of macroscopic helical assemblies. Our work clarifies the chirality evolution and transfer of inorganic nanomaterials in part without being studied previously.

A side-chain engineering strategy for constructing fluorescent dyes with direct and ultrafast self-delivery to living cells.

Organic fluorescent dyes with excellent self-delivery to living cells are always difficult to find due to the limitation of the plasma membrane having rigorous selectivity. Herein, in order to improve the permeability of dyes, we utilize a side-chain engineering strategy (SCES): adjusting the side-chain length of dyes to fine-tune the adsorption and desorption processes on the membrane–aqueous phase interfaces of the outer and inner leaflets of the plasma membrane. For this, a family of fluorescent derivatives (SPs) was prepared by functionalizing a styryl-pyridinium fluorophore with alkyl side-chains containing a different carbon number from 1 to 22. Systematic experimental investigations and simulated calculations demonstrate that the self-delivery rate of SPs with a suitable length side-chain is about 22-fold higher in SiHa cells and 76-fold higher in mesenchymal stem cells than that of unmodified SP-1, enabling cell-imaging at an ultralow loading concentration of 1 nM and deep penetration in turbid tissue and in vivo. Moreover, the SCES can even endow a membrane-impermeable fluorescent scaffold with good permeability. Further, quantitative research on the relationship between Clog P and cell permeability shows that when Clog P is in the range of 1.3–2.5, dyes possess optimal permeability. Therefore, this work not only systematically reports the effect of side-chain length on dye delivery for the first time, but also provides some ideal fluorescent probes. At the same time, it gives a suitable Clog P range for efficient cellular delivery, which can serve as a guide for designing cell-permeant dyes. In a word, all the results reveal that the SCES is an effective strategy to dramatically improve dye permeability.

Preparation and Analysis of Quantum Dots: Applications of Capillary Electrophoresis.

The implementation of quantum dots in analytical chemistry has already advanced from basic research activities to routine applications of commercially available fluorescent agents present in sophisticated assays kits. Nevertheless, a further development of new preparation and characterization methods of nanoparticles is still required to increase the sensitivity of analytical methods substantially. Thus, in many bioanalytical applications, important molecules such as DNA, proteins, and antibodies are routinely conjugated with fluorescent tags to reach even the absolute sensitivity, that is, the capability to detect a single molecule in complex matrices. Semiconductor quantum dots have already proved to be suitable components of highly luminescent tags, probes, and sensors with broad applicability in analytical chemistry. Quantum dots provide high extinction coefficients together with wide ranges of excitation wavelengths, size- and composition-tunable emissions, narrow and symmetric emission spectra, good quantum yields, relatively long size-dependent luminescence lifetime, and low photobleaching. Most of these properties are superior when compared with conventional organic fluorescent dyes. In this chapter, optimized procedures for the preparation of water-dispersed CdTe quantum dots; their coatings and conjugation reactions with antibodies, DNA, and macrocycles; and their analyses by capillary electrophoresis are described. The potential of capillary electrophoresis for fast analyses of nanoparticles, their conjugates with antibodies and immunocomplexes with targeted antigens, is demonstrated as an example.

Barbituric Acid Based Fluorogens: Synthesis, Aggregation-Induced Emission, and Protein Fibril Detection.

Fluorescent dyes, especially those emitting in the long wavelength region, are excellent candidates in the area of bioassay and bioimaging. In this work, we report a series of simple organic fluorescent dyes consisting of electron-donating aniline groups and electron-withdrawing barbituric acid groups. These dyes are very easy to construct while emitting strongly in the red region in their solid state. The photophysical properties of these dyes, such as solvatochromism and aggregation-induced emission, are systematically characterized. Afterward, the structure–property relationships of these barbituric acid based fluorogens are discussed. Finally, we demonstrate their potential applications for protein amyloid fibril detection.

Doubly Encapsulated Perylene Diimides: Effect of Molecular Encapsulation on Photophysical Properties.

Intermolecular interactions play a fundamental role on the performance of conjugated materials in organic electronic devices, as they heavily influence their optoelectronic properties. Synthetic control over the solid state properties of organic optoelectronic materials is crucial to access real life applications. Perylene diimides (PDIs) are one of the most highly studied classes of organic fluorescent dyes. In the solid state, π-π stacking suppresses their emission, limiting their use in a variety of applications. Here, we report the synthesis of a novel PDI dye that is encapsulated by four alkylene straps. X-ray crystallography indicates that intermolecular π-π stacking is completely suppressed in the crystalline state. This is further validated by the photophysical properties of the dye in both solution and solid state and supported by theoretical calculations. However, we find that the introduction of the encapsulating "arms" results in the creation of charge-transfer states which modify the excited state properties. This article demonstrates that molecular encapsulation can be used as a powerful tool to tune intermolecular interactions and thereby gain an extra level of control over the solid state properties of organic optoelectronic materials.

Physical Interactions of Biopolymers with Nanoparticles

Using fluorescence resonance energy transfer, the physical interaction of nanoparticles with biopolymers is studied by the example of the interaction of quantum dots (energy donor) with two-chain oligonucleotides labeled with a fluorescent organic dye BDP TR (energy acceptor). The high-efficiency resonance energy transfer from quantum dots to an organic dye not only makes it possible to study the dependence of the transfer efficiency on the donor-acceptor molar ratio but also represents an efficient tool for studying the interactions of oligonucleotides of different structures with nanoscale objects.

Wood powder-derived quantum dots for CeO2 photocatalytic and anti-counterfeit applications

Carbon quantum dots (CQDs or CDs) are regarded as alternatives to heavy metal quantum dots and organic fluorescent dyes. CDs show photoluminescent (PL) properties that are comparable to quantum dots and organic fluorescent dyes, but are generally eco-friendlier, less toxic and water soluble. Herein we report a novel method to prepare CDs from industrially-derived wood powder residues, which contrasts to conventional CDs synthesis that utilizes chemical agents. A solution of these wood-derived CDs was found to emit bright blue light when exposed to ultraviolet light, with a quantum yield of 47.4%. This solution was also found to significantly improve the reaction speed and efficiency of the photocatalytic degradation of CeO2. In addition, wood-based CDs performed well as a solid state, non-aqueous fluorescent and stamp-pad ink, with potential anti-fraudulent applications.

Adsorption interaction between cement hydrates minerals with fluid loss additive investigated by fluorescence technique

The mechanism of synthetic polymer fluid loss additive (FLA) was based on adsorption of FLA on the surface of hydrating cement particles and the formation of a complex network structure, resulting in decline of cement slurry filtrate. However, due to the complexity of early hydration environment of cement slurry, there was lack of effective and intuitive way to characterize the adsorption interaction between FLA and cement particles. In this study we described our quest in furthering understanding of the adsorption action of fluid loss additive on the cement minerals. By fluorescence staining technique (combined polymer additives with organic fluorescent dyes), we observed that the skin of the hydrated particles was coated with dyed polymer, which led to a successful fluid loss additive. More importantly, the analysis of the fluorescence intensity of luminous polymer in cement pore solution distinguished the respective adsorption capacity of various admixtures (dyed and undyed) on the surface of the cement particles, so as to solve the problem of the traditional method in characterizing the competitive adsorption between polymers, thus gave a convenient way to analyze the interaction mechanism between polymer admixture and cement particles from the theory and technology.