Small Dye Research Articles

Construction of Somatostatin-Based Multiphase "Core-Shell" Coacervates as Photodynamic Biomimetic Organelles.

Biomimetic coacervates have recently attracted great interest in biomedical fields, especially for drug delivery and as protocells. However, these membraneless structures are easily coalesced and poorly targetable, limiting their real biomedical applications. Here multiphase "core-shell" coacervate (CSC) constructed by dsDNA and somatostatin (SST), a 14-mer cyclopeptide is designed. The CSC shows enhanced tumor targetability through SST binding to SST receptors on the tumor cells' surface. G4 quadruplex-hemin complex can be embedded in the CSC by interaction with SST, as demonstrated by molecular simulation and isothermal titration calorimetry. The G4-hemin embedded CSC can further recruit photosensitizers such as tetracarboxyphenyl porphyrin to form the CSC-GHT composite for photodynamic therapy (PDT). As photodynamic biomimetic organelles, CSC-GHT can convert oxygen to singlet oxygen (catalyzed by the catalase-mimetic activity of G4-hemin), resulting in enhanced PDT effect, which allows the inhibition of cellular migration in vitro and tumor growth in vivo. Owing to high stability, targetability, and biosafety, the proposed CSC can recruit various cargos from small dyes to large biomacromolecules (up to 430kDa), providing promising theranostic applications.

Computational chemistry facilitates the development of second near-infrared xanthene-based dyes.

The dyes in the second near-infrared (NIR-II) region play a crucial role in advancing imaging technology. However, developing small-molecule dyes in NIR-II poses a significant bottleneck to meet the substantial demands in biological fields, which may be attributed to the lack of a rational design strategy. Herein, we designed a series of rhodamine analogs with more red-shifted emission by replacing the oxygen-bridge atom in xanthene-based dyes with -C(CH3)2, -Si(CH3)2, -SO2, and -P(O)Ph. We investigated the frontier molecular orbital, electrostatic potential surfaces, the interaction region indicator, electron-hole distribution, and absorption and emission spectrum of xanthene-based dyes using (time-dependent) density functional theory. Our results demonstrated that these designed small molecular dyes exhibit long emission wavelengths covering 1377-1809nm. We expected these findings to enable the targeted design of long-wavelength rhodamines. Geometry optimization of dyes in the ground and excited states was carried out at ω-B97XD/Def2SVP level using Gaussian 16 A03. The absorption and emission wavelengths were evaluated using 13 functional, including TPSSH, O3LYP, B3LYP*, B3LYP, PBE0, MPW1B95, PBE-1/3, PBE38, MPWB1K, MN15, BHandHLYP, ω-B97XD, and CAM-B3LYP.

Bright and Stable Cyan Fluorescent RNA Enables Multicolor RNA Imaging in Live Escherichia coli.

Fluorescent RNAs (FRs), which are RNA aptamers that bind and activate their cognate small fluorogenic dyes, have provided a particularly useful approach for imaging RNAs in live cells. Although the color palette of FRs is greatly expanded, a bright and stable cyan FR with good biocompatibility and biorthogonality with currently available FRs remains desirable but is not yet developed. Herein, the development of Myosotis is described, an RNA aptamer that emits bright cyan fluorescence upon binding a novel GFP chromophore-like fluorophore called DBT. Myosotis has a nanomolar affinity for DBT and shows a weak dependence on magnesium for folding. Further studies reveal that the Myosotis-DBT complex has a long fluorescence lifetime, good photostability, and enhance cellular brightness. It is further shown that Myosotis-DBT is biorthogonal to Pepper and Clivia FRs, allowing multiplex fluorescence imaging of RNA in live bacteria. Myosotis can also use to image mRNA in live bacteria, revealing potential coupling between mRNA translation and stability. It is believed that this cyan FR will be a useful tool for studying the functionality and mechanism of RNA underlying diverse biological processes.

Marangoni flow-guided molecular accumulation for sensitive and rapid SERS detection of phthalates

Public attention has been emerging for the prevention and assessment of healthcare risks from phthalate esters (PAEs) in daily life. For the effective detection of small environmentally hazardous materials, we suggest two major points: i) the formation of high-density hotspots and ii) the delivery of molecules to the activated plasmonic regions. In this study, we developed a method for laser-induced Marangoni flow in cotton fabric (CF) nanopillars (NPs) deposited with Au (Au/CFNPs) for the detection of PAEs by surface-enhanced Raman spectroscopy (SERS). We optimized the performance of the Au/CFNP platforms by adjusting both the maskless plasma etching time and the Au deposition thickness. Narrow (∼8 nm) and populated hotspots were achieved at an Ar plasma treatment time of 30 s and a Au thickness of 150 nm. The molecular behaviors were observed via real-time monitoring of SERS signals. Under continuous laser illumination, Marangoni radial convection flow became predominant over the outward capillary force, resulting in molecular accumulation in the detection areas. For small probe dyes and PAEs, the maximum intensities were achieved within 80 s after the injection of analytes and their signal fluctuations were estimated to have a relative standard deviation of < 10 %, which indicated sensitive and reproducible sensing. The prediction of PAEs extracted from the actual samples was investigated on the basis of a model used to quantitatively fit reference samples and the results were compared with those obtained by high-performance liquid chromatography. The results demonstrated that the combination of densified hotspots on three-dimensional porous substrates and laser-induced Marangoni flow is highly desirable for on-site early screening assays for low-quantity harmful materials present in surrounding environments.

Multipartite Fluorogenic Sensors for Monitoring Tyrosine Phosphatase Activity.

Fluorogenic substrates are essential tools for studying the activity of many enzymes including the protein tyrosine phosphatases (PTPs). Here, we have taken the first step toward the development of genetically encodable sensors for PTP activity using fluorescent and fluorogen-activating proteins. The Fluorescence-Activating and absorption Shifting Tag (FAST) is a small protein that becomes fluorescent upon binding to a small molecule dye. We demonstrate that FAST protein can be used as a sensor for PTP-mediated dephosphorylation of phosphorylated dye molecules. Phosphorylated 4-hydroxybenzylidene rhodanine (pHBR) is not able to bind to the FAST protein and induce fluorescence, but provides a sensitive assay for PTP activity, readily detecting 100 pM concentrations of PTP1B in the presence of FAST with a kcat value of 19±1 s-1 and a KM value of 93±3 μM. In addition, while phosphorylation of the C-terminal peptide of split GFP does not result in appreciable change in fluorescence of the reconstituted protein, phosphorylation of the C-terminal peptide of the split FAST protein abrogates fluorescence. Upon PTP-mediated dephosphorylation of the C-terminal peptide, the ability of the N- and C-terminal components to form a fluorescent complex with the small molecule dye is restored, leading to fluorescence.

Modulation of AIE and Intramolecular Charge Transfer of a Pyrene-Based Probe for Discriminatory Detection and Imaging of Oligomers and Amyloid Fibrils.

Oligomers and amyloid fibrils formed at different stages of protein aggregation are important biomarkers for a variety of neurodegenerative diseases including Alzheimer's and Parkinson's diseases. The development of probes for the sensitive detection of oligomeric species is important for early stage diagnosis of amyloidogenic diseases. Many small molecular dyes have been developed to probe the dynamic growth of amyloid fibrils. However, there is a lack of discriminatory detection strategies to monitor the dynamics of both oligomers and amyloid fibrils based on the differential modulation of the photophysical properties of a single dye. Here we report a pyrene-based intramolecular charge transfer (ICT) dye with large Stokes shifted red-emitting aggregation induced emission (AIE) for monitoring the dynamic populations of both oligomers and fibrils during the aggregation of hen egg white lysozyme (HEWL) protein. At the early stage of protein aggregation, the accumulation of HEWL oligomers results in a rapid and substantial increase in the red AIE intensity at 660 nm. Later, as the oligomers transform into mature fibrils, the dye exhibits a distinct photophysical change. Binding of the dye to HEWL fibrils strongly suppresses the red AIE and enhances ICT emission. This is evidenced by a gradual decrease in the AIE intensity (∼660 nm) and an increase in LE (∼490 nm) and ICT (∼540 nm) emission intensities during the later stages of protein aggregation. Thus, the dye provides simultaneous measurements of the population dynamics of both HEWL oligomers and fibrils during protein aggregation based on the discriminatory modulation of AIE and ICT of the dye. The dye also enables imaging of both HEWL oligomers and fibrils simultaneously using different emission channels in super-resolution confocal fluorescence microscopy.

Identification of Tunable, Environmentally Responsive Fluorogenic Dyes by High-Throughput Screening.

Small molecule dyes remain essential biological tools, yet only a handful of environmentally responsive fluorogenic small molecules are available for routine characterization of protein state. Here, we report the development and execution of a high throughput screen to identify compounds that increase in fluorescence in response to binding of lipophilic sites of proteins. This effort yielded two small molecules that potently indicate the presence of a range of common proteins and outperform common dyes in differential scanning fluorimetry experiments. Structure activity relationship studies revealed that these two scaffolds can be tuned both for their quantum yields and emission wavelengths. This work affords a straightforward framework for the discovery of new fluorophores and adds two fluorogenic probes to the toolbox for studying protein state.

NanoRidge filters: Fabrication strategies and performance optimization for nano-scale microfluidic particle filtration.

Filters with high throughput, minimal dead volume, and greater sensitivity to particle size are needed, which traditional benchtop filtration cannot provide. Leveraging microfabrication techniques developed by the electronics and optics industries, the filters presented here feature a unique serpentine "NanoRidge" structure, offering a continuous filtration gap spanning over three meters on a compact 4 × 14.5 mm2 footprint. This design provides more precise size filtration cut-offs and consistent flow paths compared to traditional membrane filtration systems. Despite challenges associated with glass substrate deformation impacting uniform filter gap sizes, the study provides valuable insights into the development of NanoRidge filters (NRFs) for enhancing filtration efficiency in preparatory techniques and sample analysis. This study describes the fabrication and testing of these new filter types and directly compares the performance to traditional membrane filters using the metrics of particle size cut-off (the smallest difference in particle size which can be filtered vs passed) and particle loss. The NanoRidge filters were characterized using imaging (during fabrication, post-fabrication and use, fluorescent particles captured and small molecule dye), pressure and flow measurements, and a series of particle sizes "filter or pass" studies. Particle capacity (100-250 nm) ranged from 5 × 108 to 7 × 109 in 1 ml samples at a flow rate of 100 μl/min with backpressure in the range of 1-3 Bar. The optimized fabrication procedure for the 150 nm NRF yielded a small particle recovery of 95% while also achieving a large particle filtration of 73%. High filtration efficiency was also proven in the final 60 and 80 nm NRF fabrication procedures at 96% and 91%, respectively.

Photoacoustic/ultrasound dual-modality imaging of Sentinel lymph node with carbon nanoparticles

Abstract Sentinel lymph node (SLN) biopsy is a widely used method for identifying axillary lymph node metastasis in breast cancer patients, offering an avoidance of unnecessary axillarydissections and a lower incidence of lymphedema and other complications. However, the nuclide localization has added complexity to the procedure due to its accessibility. Methylene blue and other small molecule dyes have been widely used due to their convenience, but they have been found to have lower specificity and sensitivity. Carbon nanoparticles (CNPs), as a new type of tracer, are safe and show improved specificity. However, the significant optical absorption in human tissues still renders the detection of deep lymph nodes challenging. To overcome these challenges, we developed a high-sensitivity ultrasound probe optimized for photoacoustic imaging. Based on the new probe, we constructed a high-sensitivity photoacoustic/ultrasound dual-modality imaging system. A pilot clinical study has been conducted on two patients, preliminarily verifying the system’s ability to locate SLN when combined with the CNPs.

Positive-Charge-Based Small Molecule Dyes for Gut Microbiota Fluorescent Imaging.

As one of the research hotspots in recent years, gut microbiota have been proven to be closely related to host metabolism, nutrient absorption, and immune regulation. However, there are still many urgent issues in the research of gut microbiota, such as the localization and tracking of gut microbiota. In this research, two new fluorescent probes, EF and 6F, were developed by optimizing the structure of the positron salt small molecule probe F16. In vitro labeling experiments showed that EF and 6F can quickly label Gram-positive bacteria, Staphylococcus aureus and Lactobacillus reuteri, as well as Gram-negative bacteria, Escherichia coli and Salmonella pullorum. Meanwhile, EF and 6F have little bacterial toxicity and are used at a maximum concentration of 200 μM. Compared with EF, 6F has better hydrophilicity and stronger fluorescence characteristics in aqueous solutions, making it more suitable for imaging within gut microbiota populations. The results of in vivo imaging experiments indicate that EF and 6F can label and image the intestinal microbiota colonized by the mouse intestinal mucosal epithelium without causing any damage to intestinal tissue. Compared with commercially available MitoTracker dyes and fluorescein 5-isothiocyanate (FITC) dyes, EF and 6F exhibit better biocompatibility. Therefore, the compounds EF and 6F synthesized in this study are novel small molecule probes suitable for imaging gut microbiota, providing a better probe selection for exploring complex gut microbiota.

Super-Resolution Imaging Reveals the Mechanism of Endosomal Acidification Inhibitors Against SARS-CoV-2 Infection.

In this study, super-resolution structured illumination microscope (SIM) was used to analyze molecular mechanism of endocytic acidification inhibitors in the SARS-CoV-2 pandemic, such as Chloroquine (CQ), Hydroxychloroquine (HCQ) and Bafilomycin A1 (BafA1). We fluorescently labeled the SARS-CoV-2 RBD and its receptor ACE2 protein with small molecule dyes. Utilizing SIM imaging, the real-time impact of inhibitors (BafA1, CQ, HCQ, Dynasore) on the RBD-ACE2 endocytotic process was dynamically tracked in living cells. Initially, the protein activity of RBD and ACE2 was ensured after being labeled. And then our findings revealed that these inhibitors could inhibit the internalization and degradation of RBD-ACE2 to varying degrees. Among them, 100 nM BafA1 exhibited the most satisfactory endocytotic inhibition (~63.9 %) and protein degradation inhibition (~97.7 %). And it could inhibit the fusion between endocytic vesicles in the living cells. Additionally, Dynasore, a widely recognized dynein inhibitor, also demonstrated cell acidification inhibition effects. Together, these inhibitors collectively hinder SARS-CoV-2 infection by inhibiting both the viral internalization and RNA release. The comprehensive evaluation of pharmacological mechanisms through super-resolution fluorescence imaging has laid a crucial theoretical foundation for the development of potential drugs to treat COVID-19.

Coupling Nanoscale Precision with Multiscale Imaging: A Multifunctional Near-Infrared Dye for the Brain.

Live imaging of primary neural cells is crucial for monitoring neuronal activity, especially multiscale and multifunctional imaging that offers excellent biocompatibility. Multiscale imaging can provide insights into cellular structure and function from the nanoscale to the millimeter scale. Multifunctional imaging can monitor different activities in the brain. However, this remains a challenge because of the lack of dyes with a high signal-to-background ratio, water solubility, and multiscale and multifunctional imaging capabilities. In this study, we present a neural dye with near-infrared (NIR) emissions (>700 nm) that enables ultrafast staining (in less than 1 min) for the imaging of primary neurons. This dye not only enables multiscale neural live-cell imaging from vesicles in neurites, neural membranes, and single neurons to the whole brain but also facilitates multifunctional imaging, such as the monitoring and quantifying of synaptic vesicles and the changes in membrane potential. We also explore the potential of this NIR neural dye for staining brain slices and live brains. The NIR neural dye exhibits superior binding with neural membranes compared to commercial dyes, thereby achieving multiscale and multifunctional brain neuroimaging. In conclusion, our findings introduce a significant breakthrough in neuroimaging dyes by developing a category of small molecular dyes.

Double-bridged N–B←N bipyridyl-based airfoil-shaped small molecule dyes: π-bridge regulation on photovoltaic performance

The double-bridged N–B←N bipyridine unit (BNBP) with boron-nitrogen covalent bond and coordination bond can reduce the electronic energy level of the π-conjugated system, resulting in a reduction of band gap and a red shift in the absorption spectrum. Embedding the BNBP building block into organic optoelectronic materials provides a novel possibility in molecule design. In this paper, under the guidance of density functional theory (DFT) calculations, a series of 3D “airfoil-shaped” small molecule donors (SMDs) have been designed and successfully synthesized through Sonogashira, Suzuki and Stille coupling reactions. BNBP is adopted as the central electron-withdrawing unit and strong electron-rich triphenylamine (TPA) as the terminal group. The novel “airfoil-shaped” structure plays a supporting role in reducing the carrier recombination in the active layer. It reveals that molecular design engineering can achieve the expected results by the following steps: Firstly, the introduction of electron-rich alkoxy groups into the end of TPA through side chain engineering helps to improve the solubility and film-forming properties of the materials. Then, the π-bridge regulation not only extends the conjugate structure, but also plays a crucial role in the regulation of photovoltaic properties. It is worth noting that the compound BNBP(3TTPA)2 was synthesized by introducing thiophene oligomer into BNBP-based SMDs for the first time. Finally, the compound BNBP(EDPPTPA)2 was constructed by combining BNBP with DPP dye unit. Its narrow band gap is only 1.36 eV, which is one of the narrowest band gaps in small molecular organic-boron materials. In particular, compared with the starting compound BNBP(ETPA)2, the synergistic effects of DPP and BNBP broaden the edge absorption wavelength to 814 nm, resulting in a red shift of 170 nm. The power conversion efficiency (PCE) of 4.48 % is obtained in BNBP(EDPPTPA)2-based bulk heterojunction (BHJ) organic solar cells (OSCs), which is more than twice that of reference compound BNBP(ETPA)2. This work provides important references for the future development of 3D BNBP-based organic-boron SMDs and the structural regulation of materials through molecular engineering.

Electrochemical degradation of small molecule dyes by TiO2-decorated polyacrylonitrile nanofiber membranes with superior properties

To optimize the uniform dispersion of nanoparticles and enhance their catalytic degradation effect on small molecule dyes in textile wastewater, titanium dioxide (TiO2) was deposited on polyacrylonitrile (PAN) fiber membranes using magnetron sputtering technology, and TiO2/PAN nanofiber membrane (NFM) with low content and high catalytic efficiency was successfully prepared. Through membrane separation coupling electrocatalysis, the composite membranes exhibited outstanding performance in removing Congo red (CR), Sunset yellow (SY), Methyl orange (MO), and Methylene blue (MB) dyes, with removal rates of 99.5%, 98.8%, 98.5%, and 87.2% respectively. Even after continuous catalysis for 10 h, the composite membranes maintained a high dye removal rate, demonstrating their remarkable stability and water treatment capabilities. Furthermore, TiO2/PAN NFMs displayed high pure water permeance, dye permeance, and mechanical strength. Specifically, M20 (magnetron sputtering time of 20 min) showed water permeance and dye (SY) permeance values of 1944 L·m−2·h−1·bar−1 and 816 L·m−2·h−1·bar−1, respectively, and indicate superior mechanical and electrochemical properties. During the electrocatalytic process, TiO2/PAN NFMs generated reactive oxygen species (ROS) on the surface, imparting a self-cleaning property that extended the service life of the membranes and offered an effective solution for textile wastewater treatment.

Machine learning-assisted fluorescence/fluorescence colorimetric sensor array for discriminating amyloid fibrils

Misfolding and aggregation of proteins often lead to the development of diseases, and amyloid has gained widespread attention as a biomarker for a variety of diseases. In this study, we developed a fluorescence/fluorescence colorimetric dual-mode sensor array for the detection of amyloid fibrils using several commercially available organic small molecular dyes and alkaloids, including Thioflavin T, Congo Red, 8-anilino-1-naphthalenesulfonic acid, Safranine T, berberine and coptisine, as the elements. Herein, the array could not only use the fluorescence intensities change before and after protein interaction as a pattern recognition signal, but also read the ΔR/ΔG/ΔB values of the photos taken in the UV dark box on a smartphone-based platform, which converted the chromaticity information into intuitive data. Five studied amyloid fibrils, i.e. insulin, lysozyme, bovine serum albumin, amyloid-β 42 and α-synuclein fibrils, were properly distinguished with data processing assisted by machine learning algorithms, i.e. linear discriminant analysis, principal component analysis and hierarchical cluster analysis. After reducing the number of elements by principal component analysis, a simplified array quantified individual amyloid fibrils at 0.05–5 μM and 0.5–10 μM with fluorescence and fluorescence colorimetric signals, respectively, and successfully identified 25 unknown samples with high accuracy in diluted human plasma matrix and artificial cerebrospinal fluid. The array had good selectivity and sensitivity, providing a simple and inexpensive method for amyloid discrimination.

Microfluidic production, stability and loading of synthetic giant unilamellar vesicles

In advanced drug delivery, versatile liposomal formulations are commonly employed for safer and more accurate therapies. Here we report a method that allows a straightforward production of synthetic monodisperse (~ 100 μm) giant unilamellar vesicles (GUVs) using a microfluidic system. The stability analysis based on the microscopy imaging showed that at ambient conditions the produced GUVs had a half-life of 61 ± 2 h. However, it was observed that ~ 90% of the calcein dye that was loaded into GUVs was transported into a surrounding medium in 24 h, thus indicating that the GUVs may release these small dye molecules without distinguishable membrane disruption. We further demonstrated the feasibility of our method by loading GUVs with larger and very different cargo objects; small soluble fluorescent proteins and larger magnetic microparticles in a suspension. Compared to previously reported microfluidics-based production techniques, the obtained results indicate that our simplified method could be equally harnessed in creating GUVs with less cost, effort and time, which could further benefit studying closed membrane systems.

Systematic Structural Modification of Squaraine Dye for Near-Infrared Window One and Two Multiplexed In Vivo Imaging and Photothermal Therapy.

Due to the wide application of reporter gene-related visible/NIR-I bioluminescent imaging, multiplexed fluorescence imaging across visible/NIR-I/NIR-II has excellent potential in biomedical research. However, in vivo multiplexed imaging applications across those regions have rarely been reported due to the lack of proper fluorophores. Herein, nine squaraine dyes, which exhibit diverse adsorption and emission wavelengths, were synthesized. Among them, water-soluble SQ 710-5k and SQ 905 were found to have significant absorption differences, which allowed the tumor and lymph nodes to be identified. Then, for the first time, six-channel multiplexed fluorescence imaging across visible/NIR-I/II was achieved by coordination with reporter gene-related bioluminescent phosphors. Additional research revealed that SQ 710-5k exhibited higher-quality blood vessels and tumor imaging in NIR-II. H-aggregates SQ 905 demonstrated a high photothermal conversion efficiency for photothermal therapy. This study proposed an approach to creating small molecular dyes that coordinate with reporter gene-related bioluminescent phosphors for six-color fluorescence imaging.

Design and synthesis of novel A-D-A type small molecule dyes

Abstract This article introduces two novel A-D-A type small molecules. Compared to traditional methylene-based dye materials, these compounds exhibit enhanced conjugation and improved stability. The newly studied A-D-A type small molecules in this research possess narrow bandgaps, broad absorption in the visible light range, strong fluorescence, and good solubility.

CO2-Sourced Polymer Dyes for Dual Information Encryption.

Large amounts of small molecule dyes leak into the ecosystems annually in harmful and unsustainable ways. Polymer dyes have attracted much attention because of their high migration resistance, excellent stability, and minimized leakage. However, the complex synthesis process, high cost, and poor degradability hinder their widespread application. Herein, green and sustainable polymer dyes are prepared using natural dye quercetin (Qc) and CO2 through a one-step process. The CO2-sourced polymer dyes show strong migration resistance, high stability, and can be degraded on demand. Additionally, the CO2-sourced polymer dyes showed unique responses to Zn2+, leading to significantly enhanced fluorescence, highlighting their potential for information encryption/decryption. The CO2-sourced polymer dyes can solve the environmental hazards caused by small molecule dye leakage and promote the carbon cycle process. Meanwhile, the one-step synthesis process is expected to achieve sustainable and widespread utilization of CO2-sourced polymer dyes.

Simple cyclic chalcone dye with multiple optical functions: Piezochromism and lysosomes staining

Both artificially synthesized and naturally occurring cyclic chalcones have been widely studied for their excellent biological activities. However, research on its photophysical properties is still limited. In the present study, we designed and synthesized a small molecule fluorescent dye based on the ICT effect, using dimethylamino as the electron-donating group and carbonyl as the electron withdrawing group, and investigated its photophysical properties in depth. Although YB is a simple small molecule, it exhibits significant piezochromic properties. The fluorescence of YB can change from green to yellow through grinding. After solvent fumigation, the fluorescence reverts to green. Furthermore, YB was used successfully in the lysosomal targeting. This study expands the research on the photophysical properties of cyclic chalcone and give richness to application of cyclic chalcone compounds.