3-Ethoxysalicylaldimine-based symmetrical azine-linked luminogen exhibiting ESIPT and bright orange colour AIE behaviour with live cell bioimaging application.

Herein, we describe the design and development of a new cell-permeable aggregation-induced emission (AIE) active 3-ethoxysalicylaldimine-based symmetrical azine molecule HDBE. The synthesized compound underwent comprehensive investigation of different spectroscopic methods, like NMR, mass and single crystal X-ray diffraction analysis. The fluorophore HDBE exhibited the bright orange colour AIE behaviour in THF-H2O mixture. The drastic enhancement of emission was achieved upon adding the water to the THF solution of HDBE, with a concentration of 90%. Along with the dynamic light scattering (DLS) and quantum yield measurements, the formation of aggregates was also verified by scanning electron microscopy (SEM) and transmission electron microscopy (TEM) analysis. Further, HDBE demonstrated excited state intramolecular proton transfer (ESIPT) characteristics in different polarity of solvents, which was corroborated by absorption, emission and lifetime spectroscopical investigations. The detailed scrutiny of X-ray structure of HDBE displayed the two strong intramolecular hydrogen bonding interactions, while solid-state fluorescent spectra showed dual emission that corresponds to enol and keto form confirming the ESIPT feature. Further, the synthesized AIE molecule was non-toxic and cell-permeable, making it easy to label as a biomarker in live HeLa cells via fluorescent bioimaging. These studies offer a quick and easy way to develop both AIE and ESIPT-coupled molecules for live cell bioimaging applications.

Including the rare cubane cluster cobalt coordination polymer as the fluorescent sensing material for selectively and sensitively detecting the nitrofurantoin antibiotic.

More and more attention has been paid to food safety. Due to the overuse and misuse of antibiotics, the problem of antibiotic residues in animal food is one of the important challenges to ensure food safety. The development of a feasible strategy to detect antibiotic residues in animal food has become desirable. In this paper, we creatively synthesize a water-stable fluorescence sensing material, namely, Co(Ⅱ)-Coordination polymer [Co2(CA) (L)0.5 (H2O)3] n (L = 1,4-bis(imidazole-1-ylmethyl) benzene, CA= Citric acid). The single crystal X-ray diffraction shows that it crystallizes in tetragonal space group I-4. It is worth mentioning that there exists the rare Co4(µ3-O)4 cubane cluster structure and Co8 cluster units. Those adjacent Co8 cluster units are connected into an infinite two-dimensional net structure by four flexible bridged L ligands. Finally, the Co(Ⅱ)-Coordination polymer (CP) further develops into the three-dimensional supramolecular structure via the hydrogen bonds of O-H⋯O and C-H⋯O. It could selectively detect the antibiotic-nitrofurantoin (NFT) residue by way of fluorescence quenching, Co-CP for the detection of NFT shows broad linearity from 0 to 200 µM, with a detection limit of 0.13 µM and strong anti-interference ability. It is used to detect the NFT residual of tap water and milk with a spiked recovery of 86.35-112.47 %.

Conceptually innovative fluorophores for functional bioimaging.

Fluorophore chemistry is at the forefront of bioimaging, revolutionizing the visualization of biological processes with unparalleled precision. From the serendipitous discovery of mauveine in 1856 to cutting-edge fluorophore engineering, this field has undergone transformative evolution. Today, the synergy of chemistry, biology, and imaging technologies has produced diverse, specialized fluorophores that enhance brightness, photostability, and targeting capabilities. This review delves into the history and innovation of fluorescent probes, showcasing their pivotal role in advancing our understanding of cellular dynamics and disease mechanisms. We highlight groundbreaking molecules and their applications, envisioning future breakthroughs that promise to redefine biomedical research and diagnostics.

Fluorescent labeling of micro/nanoplastics for biological applications with a focus on "true-to-life" tracking.

The increased environmental presence of micro-/nanoplastics (MNPLs) and the potential health risks associated with their exposure classify them as environmental pollutants with special environmental and health concerns. Consequently, there is an urgent need to investigate the potential risks associated with secondary MNPLs. In this context, using "true-to-life" MNPLs, resulting from the laboratory degradation of plastic goods, may be a sound approach. These non-commercial secondary MNPLs must be labeled to track their presence/journeys inside cells or organisms. Because the cell internalization of MNPLs is commonly analyzed using fluorescence techniques, the use of fluorescent dyes may be a sound method to label them. Five different compounds comprising two chemical dyes (Nile Red and Rhodamine-B), one optical brightener (Opticol), and two industrial dyes (Amarillo Luminoso and iDye PolyPink) were tested to determine their potential for such applications. Using commercial standards of polystyrene nanoplastics (PSNPLs) with an average size of 170 nm, different characteristics of the selected dyes such as the absence of impact on cell viability, specificity for plastic staining, no leaching, and lack of interference with other fluorochromes were analyzed. Based on the overall data obtained in the wide battery of assays performed, iDye PolyPink exhibited the most advantages, with respect to the other compounds, and was selected to effectively label "true-to-life" MNPLs. These advantages were confirmed using a proposed protocol, and labeling titanium-doped PETNPLs (obtained from the degradation of milk PET plastic bottles), as an example of "true-to-life" secondary NPLs. These results confirmed the usefulness of iDye PolyPink for labeling MNPLs and detecting cell internalization.

Exploring the Properties of Unilamellar Vesicle Bilayers Formed by Ionic Liquid Surfactants for Future Applications in Nanomedicine.

Two ionic liquids (ILs) with amphiphilic properties composed of 1-butyl-3-methylimidazolium dioctylsulfosuccinate (bmim-AOT) and 1-hexyl-3-methylimidazolium dioctylsulfosuccinate (hmim-AOT) form unilamellar vesicles spontaneously simply by dissolving the IL-like surfactant in water. These novel vesicles were characterized using two different and highly sensitive fluorescent probes: 6-propionyl-2-(dimethylaminonaphthalene) (PRODAN) and trans-4-[4-(dimethylamino)-styryl]-1-methylpyridinium iodide (HC). These fluorescent probes provide information about the physicochemical properties of the bilayer, such as micropolarity, microviscosity, and electron-donor capacity. In addition, the biocompatibility of these vesicles with the blood medium was evaluated, and their toxicity was determined using Dictyostelium discoideum amoebas. First, using PRODAN and HC, it was found that the bilayer composition and the chemical structure of the ions at the interface produced differences between both amphiphiles, making the vesicles different. Thus, the bilayer of hmim-AOT vesicles is less polar, more rigid, and has a lower electron-donor capacity than those made by bmim-AOT. Finally, the results obtained from the hemolysis studies and the growth behavior of unicellular amoebas, particularly utilizing the D. discoideum assay, showed that both vesicular systems do not produce toxic effects up to a concentration of 0.02 mg/mL. This elegant assay, devoid of animal usage, highlights the potential of these newly organized systems for the delivery of drugs and bioactive molecules of different polarities.

Development of a "turn off" fluorescent molecularly imprinted nanoparticle-based sensor for selective captopril quantification in synthetic urine and wastewater samples.

The combination of silica nanoparticles with fluorescent molecularly imprinted polymers (Si-FMIPs) prepared by a one-pot sol-gel synthesis method to act as chemical sensors for the selective and sensitive determination of captopril is described. Several analytical parameters were optimized, including reagent ratio, solvent, concentration of Si-FMIP solutions, and contact time. Fourier-transform infrared spectroscopy (FT-IR), transmission electron microscopy (TEM), and the ninhydrin assay were used for characterization. The selectivity was evaluated against molecules belonging to other drug classes, such as fluoroquinolones, nonacid nonopioids, benzothiadiazine, alpha amino acids, and nitroimidazoles. Under optimized conditions, the Si-FMIP-based sensor exhibited a working range of 1-15 µM, with a limit of detection (LOD) of 0.7 µM, repeatability of 6.4% (n = 10), and suitable recovery values at three concentration levels (98.5% (1.5 µM), 99.9% (3.5 µM), and 99.2% (7.5 µM)) for wastewater samples. The sensor provided a working range of 0.5-15 µM for synthetic urine samples, with an LOD of 0.4 µM and a repeatability of 7.4% (n = 10) and recovery values of 93.7%, 92.9%, and 98.0% for 1.0 µM, 3.5 µM, and 10 µM, respectively. In conclusion, our single-vessel synthesis approach for Si-FMIPs proved to be highly effective for the selective determination of captopril in wastewater and synthetic urine samples.

Production of Peroxymonocarbonate by Steady-State Micromolar H2O2 and Activated Macrophages in the Presence of CO2/HCO3- Evidenced by Boronate Probes.

Peroxymonocarbonate (HCO4-/HOOCO2-) is produced by the reversible reaction of CO2/HCO3- with H2O2 (K = 0.33 M-1, pH 7.0). Although produced in low yields at physiological pHs and H2O2 and CO2/HCO3- concentrations, HCO4- oxidizes most nucleophiles with rate constants 10 to 100 times higher than those of H2O2. Boronate probes are known examples because HCO4- reacts with coumarin-7-boronic acid pinacolate ester (CBE) with a rate constant that is approximately 100 times higher than that of H2O2 and the same holds for fluorescein-boronate (Fl-B) as reported here. Therefore, we tested whether boronate probes could provide evidence for HCO4- formation under biologically relevant conditions. Glucose/glucose oxidase/catalase were adjusted to produce low steady-state H2O2 concentrations (2-18 µM) in Pi buffer at pH 7.4 and 37 °C. Then, CBE (100 µM) was added and fluorescence increase was monitored with time. The results showed that each steady-state H2O2 concentration reacted more rapidly (∼30%) in the presence of CO2/HCO3- (25 mM) than in its absence, and the data permitted the calculation of consistent rate constants. Also, RAW 264.7 macrophages were activated with phorbol 12-myristate 13-acetate (PMA) (1 µg/mL) at pH 7.4 and 37 °C to produce a time-dependent H2O2 concentration (8.0 ± 2.5 µM after 60 min). The media contained 0, 21.6, or 42.2 mM HCO3- equilibrated with 0, 5, or 10% CO2, respectively. In the presence of CBE or Fl-B (30 µM), a time-dependent increase in the fluorescence of the bulk solution was observed, which was higher in the presence of CO2/HCO3- in a concentration-dependent manner. The Fl-B samples were also examined by fluorescence microscopy. Our results demonstrated that mammalian cells produce HCO4- and boronate probes can evidence and distinguish it from H2O2 under biologically relevant concentrations of H2O2 and CO2/HCO3-.

Cell-Based Thallium-Influx Fluorescence Assay for Kv10.1 Channels.

The development of cell-based fluorescent assays has resulted in an incredible tool for searching new ion channels' modulators with a biophysical and clinical profile. Among all the ion channels, potassium (K+)-permeable channels represent the most diverse and relevant for cell function, making them attractive targets for drug discovery. Some of the cell-based assays for K+ channels take advantage of a thallium-sensitive dye whose fluorescence increased upon the binding of thallium (Tl+), an ion able to move through K+ channels. We optimize the FLIPR Potassium Assay Kit based on thallium influx to measure the Kv10.1 activity.

Molecular two-point recognition of fructosyl valine and fructosyl glycyl histidine in water by fluorescent Zn(II)-terpyridine complexes bearing boronic acids.

Selective recognition of fructosyl amino acids in water by arylboronic acid-based receptors is a central field of modern supramolecular chemistry that impacts biological and medicinal chemistry. Fructosyl valine (FV) and fructosyl glycyl histidine (FGH) occur as N-terminal moieties of human glycated hemoglobin; therefore, the molecular design of biomimetic receptors is an attractive, but very challenging goal. Herein, we report three novel cationic Zn-terpyridine complexes bearing a fluorescent N-quinolinium nucleus covalently linked to three different isomers of strongly acidified phenylboronic acids (ortho-, 2Zn; meta-, 3Zn and para-, 4Zn) for the optical recognition of FV, FGH and comparative analytes (D-fructose, Gly, Val and His) in pure water at physiological pH. The complexes were designed to act as fluorescent receptors using a cooperative action of boric acid and a metal chelate. Complex 3Zn was found to display the most acidic -B(OH)2 group (pKa = 6.98) and exceptionally tight affinity for FV (K = 1.43 × 105 M-1) with a strong quenching analytical response in the micromolar concentration range. The addition of fructose and the other amino acids only induced moderate optical changes. On the basis of several spectroscopic tools (1H, 11B NMR, UV-Vis, and fluorescence titrations), ESI mass spectrometry, X-ray crystal structure, and DFT calculations, the interaction mode between 3Zn and FV is proposed in a 1 : 1 model through a cooperative two-point recognition involving a sp3 boronate-diol esterification with simultaneous coordination bonding of the carboxylate group of Val to the Zn atom. Fluorescence quenching is attributed to a static complexation photoinduced electron transfer mechanism as evidenced by lifetime experiments. The addition of FGH to 3Zn notably enhanced its emission intensity with micromolar affinity, but with a lower apparent binding constant than that observed for FV. FGH interacts with 3Zn through boronate-diol complexation and coordination of the imidazole ring of His. DFT-optimized structures of complexes 3Zn-FV and 3Zn-FGH show a picture of binding which shows that the Zn-complex has a suitable (B⋯Zn) distance to the two-point recognition with these analytes. Molecular recognition of fructosyl amino acids by transition-metal-based receptors has not been explored until now.

Exploring mitochondrial targeting: an innovative fluorescent probe reveals Nernstian potential and partitioning combination.

This study introduces a paradigm-shifting approach to optimize mitochondrial targeting. Employing a new fluorescent probe strategy, we unravel a combined influence of both Nernst potential (Ψ) and partitioning (P) contributions. Through the synthesis of new benz[e]indolinium-derived probes, our findings redefine the landscape of mitochondrial localization by optimizing the efficacy of mitochondrial probe retention in primary cortical neurons undergoing normoxia and oxygen-glucose deprivation. This methodology not only advances our understanding of subcellular dynamics, but also holds promise for transformative applications in biomedical research and therapeutic development.

Optical recordings of organellar membrane potentials and the components of membrane conductance in lysosomes.

The eukaryotic cell is highly compartmentalized with organelles. Owing to their function in transporting metabolites, metabolic intermediates and byproducts of metabolic activity, organelles are important players in the orchestration of cellular function. Recent advances in optical methods for interrogating the different aspects of organellar activity promise to revolutionize our ability to dissect cellular processes with unprecedented detail. The transport activity of organelles is usually coupled to the transport of charged species; therefore, it is not only associated with the metabolic landscape but also entangled with membrane potentials. In this context, the targeted expression of fluorescent probes for interrogating organellar membrane potential (Ψorg) emerges as a powerful approach, offering less-invasive conditions and technical simplicity to interrogate cellular signalling and metabolism. Different research groups have made remarkable progress in adapting a variety of optical methods for measuring and monitoring Ψorg. These approaches include using potentiometric dyes, genetically encoded voltage indicators, hybrid fluorescence resonance energy transfer sensors and photoinduced electron transfer systems. These studies have provided consistent values for the resting potential of single-membrane organelles, such as lysosomes, the Golgi and the endoplasmic reticulum. We can foresee the use of dynamic measurements of Ψorg to study fundamental problems in organellar physiology that are linked to serious cellular disorders. Here, we present an overview of the available techniques, a survey of the resting membrane potential of internal membranes and, finally, an open-source mathematical model useful to interpret and interrogate membrane-bound structures of small volume by using the lysosome as an example.

A pH-Sensitive Fluorescent Chemosensor Turn-On Based in a Salen Iron (III) Complex: Synthesis, Photophysical Properties, and Live-Cell Imaging Application.

pH regulation is essential to allow normal cell function, and their imbalance is associated with different pathologic situations, including cancer. In this study, we present the synthesis of 2-(((2-aminoethyl)imino)methyl)phenol (HL1) and the iron (III) complex (Fe(L1)2Br, (C1)), confirmed by X-ray diffraction analysis. The absorption and emission properties of complex C1 were assessed in the presence and absence of different physiologically relevant analytes, finding a fluorescent turn-on when OH- was added. So, we determined the limit of detection (LOD = 3.97 × 10-9 M), stoichiometry (1:1), and association constant (Kas = 5.86 × 103 M-1). Using DFT calculations, we proposed a spontaneous decomposition mechanism for C1. After characterization, complex C1 was evaluated as an intracellular pH chemosensor on the human primary gastric adenocarcinoma (AGS) and non-tumoral gastric epithelia (GES-1) cell lines, finding fluorescent signal activation in the latter when compared to AGS cells due to the lower intracellular pH of AGS cells caused by the increased metabolic rate. However, when complex C1 was used on metastatic cancer cell lines (MKN-45 and MKN-74), a fluorescent turn-on was observed in both cell lines because the intracellular lactate amount increased. Our results could provide insights about the application of complex C1 as a metabolic probe to be used in cancer cell imaging.

A fluorescent glyconanoprobe based on quantum dots and thiolated glucose: Applications in monolayers and spheroids of cancer cells.

The differential energy metabolism of cancer cells has stimulated the development of tools that can be applied to better understand the complex biological interaction involved in the uptake of glucose analogs at the cellular level in this disease. Herein, we explored the outstanding optical properties of quantum dots (QDs) to develop a new fluorescent glyconanoprobe using the 1-thio-ß-d-glucose (Glc). Then, monolayers and spheroids of HeLa cells were applied to probe the biological interaction with the conjugate through fluorescence techniques. Spheroids have been gaining prominence for better mimicking the tumor microenvironment. The Glc-QDs conjugate was prepared by a facile and direct procedure based on the affinity of the Glc thiol group by the QD semiconductor surface. The conjugation was evaluated and confirmed by Zeta potential (ζ) measurements, FTIR spectroscopy, and fluorescence correlation spectroscopy (FCS). Moreover, a biological assay using Candida albicans yeasts coated with concanavalin A, by exploring the lectin-carbohydrate affinity, was also developed to further confirm the conjugation, which corroborated the previous analyses. The hanging drop method was used to prepare the spheroids. The fluorescence microscopy analyses indicated an intracellular labeling by the glyconanoprobe, in both cell culture models. Flow cytometry assays revealed effective uptake of the conjugate (above ca. 76%), even by cells cultivated as spheroids, applying short incubation time. Therefore, a new fluorescent glyconanoprobe was developed, which showed potential to be applied for investigating mechanisms involved in the uptake of glucose analogs, both by simpler and complex cancer biological models, as monolayers and spheroids.

Lemon-derived carbon dots as antioxidant and light emitter in fluorescent films applied to nanothermometry.

The design of luminescent nanomaterials for the development of nanothermometers with high sensitivity and free of potentially toxic metals has developed in several fields, such as optoelectronics, sensors, and bioimaging. In addition, luminescent nanothermometers have advantages related to non-invasive measurement, with their wide detection range and high spatial resolution at the nano/microscale. Our study is the first, to our knowledge, to demonstrate a detailed study of a fluorescent film (Film-L) thermal sensor based on carbon dots derived from lemon bagasse extract (CD-L). The CD-L properties were explored as an antioxidant agent; their cytotoxicity was evaluated by using a human non-tumoral skin fibroblast (HFF-1) cell line from an MTT assay. The CD-L were characterized by HRTEM, DLS, FTIR, UV-VIS, and fluorescence spectroscopy. These confirmed their particle size distribution below 10 nm, graphitic structure in the core and surface organic groups, and strong blue emission. The CD-L showed cytocompatibility behavior and scavenging potential reactive species of biological importance: O2•- and HOCl, with IC50 of 276.8 ± 4.0 and 21.6 ± 0.7, respectively. The Film-L emission intensities (I425 nm) are temperature-dependent in the 298 to 333 K range. The Film-L luminescent thermometer shows a maximum relative thermal sensitivity of 2.69 % K-1 at 333 K.

Fluorescent Probe for in Vivo Partitioning into Dynamic Lipid Droplets Enables Monitoring of Water Permeability-Induced Edema.

Lipid droplets (LDs) are intracellular organelles found in most cell types from adipocytes to cancer cells. Although recent investigations have implicated LDs in numerous diseases, the current available methods to monitor them in vertebrate models rely on static imaging using fluorescent dyes, limiting the investigation of their rapid in vivo dynamics. Here, we report a fluorophore chemistry approach to enable in vivo LD dynamic monitoring using a Nernstian partitioning mechanism. Interestingly, the effect of atorvastatin and osmotic treatments toward LDs revealed an unprecedented dynamic enhancement. Then, using a designed molecular probe with an optimized response to hydration and LD dynamics applied to Zebrafish developing pericardial and yolk-sac edema, which represents a tractable model of a human cardiovascular disease, we also provide a unique dual method to detect disease evolution and recovery.

Study of the pH effect on the optical and morphological properties of S, N self-doped carbon dots applied as fluorescent anti-counterfeiting ink and pH sensor.

The pH value is an important parameter as it is part of several processes, whether environmental or biological. In this report, S, N self-doped carbon dots (CDs) were synthesized by a simple hydrothermal method using cysteine (cys) and citric acid as precursors for a detailed investigation of size, morphological, photoluminescent, and structural changes at different pH values and its use as pH sensor and fluorescent ink. The fluorescence intensity of cys-CDs was dependent on the pH, presenting a linear relationship with pH values in the range of 2.0-9.0. Using spectroscopic techniques, a mechanism for the pH-dependent fluorescence is proposed, based on the aggregation of cys-CDs and also protonation/deprotonation of surface functional groups that change the excited state. The cys-CDs were found to be efficient as fluorescent pH sensors using real samples (distilled water and tap water). Furthermore, the pH changes in cys-CDs can be used for the visual enhancement of anti-counterfeiting technologies. Thus, the results of this study show that cys-CDs can act as an efficient and pH sensitive fluorescent sensor, which can be used to measure the pH value of water samples, due to its high fluorescence intensity, and can be applied successfully as a fluorescent ink.

Red fluorescent benzothiadiazole derivative loaded in different nanoformulations: Optical properties and their use in bio-imaging.

Fluorophores with optimized nonlinear optical properties have become prominent as contrast labels in laser scanning microscopy (LSM). The purpose of this work is to report on a novel benzothiadiazole derivative, namely 4,7-bis(5-((9,9-dioctyl-9H-fluoren-2-yl)ethynyl)thiophen-2-yl)benzo[c][1,2,5]thiadiazole (EFBT) and its optical performance when it is loaded into organic nanostructures intended as labels for LSM. Four different nanostructured labels were prepared: i) EFBT-loaded silica nanoparticles (SiNPs); ii) folate-bioconjugated SiNPs (SiNPs-FA); iii) EFBT-loaded PEGylated nanoparticles (NPs-PEG); and iv) EFBT-loaded folate-terminated PEGylated nanoparticles (NPs-PEG-FA). All these nanostructures are reported through a comparative study of their linear and nonlinear optical properties, including their performance as exogenous label agents in the cervical cancer cell line HeLa. This assessment of the performance of a specific fluorophore loaded into different nanostructured matrices (labels), and fairly compared under the same characterization conditions, including the LSM settings, is less common while previous reports had focused in comparing silica and PEGylated nanoparticles but loaded with different fluorophores. The results show that the internal molecular organization into each type of organic nanostructure impacted differently the properties of EFBT, where the silica matrix tend to preserve the optical performance of the fluorophore by preventing intermolecular interactions; in contrast, PEGylated nanoparticles favored molecular interactions and introduced non-radiative decay channels that degrades drastically the optical performance. Nevertheless, the use of functionalized ends entities produced a better cellular label uptake with PEGylated that with silica nanoparticles. In overall, the NPs-PEG-FA label produced the best HeLa imaging.

Simple and cheap preparation of fluorescence paper sensor based in carbon dot for visual detection of chloramphenicol.

Carbon dots (CDs) are nanometer-scale particles produced from carbon sources that exhibit fluorescence emission. The present work presents the synthesis and characterization of CDs, as well as the sensing studies for the determination of chloramphenicol (CAP). CAP is an antibiotic used in human medicine and agriculture, and its indiscriminate use and inappropriate disposal have caused damage to human health and the environment. The carbonaceous precursor used in the synthesis of CDs was 3,4-diaminobenzoic acid (3,4-DABA) through the hydrothermal method via domestic microwave irradiation. The first synthesis procedure was carried out in the presence of water/ethanol (a-CDs) and the second in the presence of 1 mol/L sodium hydroxide/ethanol (b-CDs). The CDs were initially characterized in terms of spectroscopic properties in the ultraviolet and visible region (UV-visible), Fourier-transform infrared (FTIR) spectra, Raman spectroscopy, and fluorescence emission spectroscopy. Sensing studies for the antibiotic C were performed by fluorescence suppression in the presence of a- and b-CDs, as well as the precursor 3,4-DABA. The a- and b-CDs presented similar values of linear range 0.00080-0.0050 mg/ml and limit of detection (LOD) = 0.00030 mg/ml (0.30 ppm) for CAP. Then, a- and b-CDs were embedded in Whatman and Mellita® filter paper, and CAP sensing was evaluated through UV light excitation.

Single-Fluorophore Indicator to Explore Cellular and Sub-cellular Lactate Dynamics.

Lactate is an energy substrate and an intercellular signal, which can be monitored in intact cells with the genetically encoded FRET indicator Laconic. However, the structural complexity, need for sophisticated equipment, and relatively small fluorescent change limit the use of FRET indicators for subcellular targeting and development of high-throughput screening methodologies. Using the bacterial periplasmic binding protein TTHA0766 from Thermus thermophilus, we have now developed a single-fluorophore indicator for lactate, CanlonicSF. This indicator exhibits a maximal fluorescence change of 200% and a KD of ∼300 µM. The fluorescence is not affected by other monocarboxylates. The lactate indicator was not significantly affected by Ca2+ at the physiological concentrations prevailing in the cytosol, endoplasmic reticulum, and extracellular space, but was affected by Ca2+ in the low micromolar range. Targeting the indicator to the endoplasmic reticulum revealed for the first time sub-cellular lactate dynamics. Its improved lactate-induced fluorescence response permitted the development of a multiwell plate assay to screen for inhibitors of the monocarboxylate transporters MCTs, a pharmaceutical target for cancer and inflammation. The functionality of the indicator in living tissue was demonstrated in the brain of Drosophila melanogaster larvae. CanlonicSF is well suited to explore lactate dynamics with sub-cellular resolution in intact systems.

Doped Carbon Quantum Dots/PVA Nanocomposite as a Platform to Sense Nitrite Ions in Meat.

A sensor device based on doped-carbon quantum dots is proposed herein for detection of nitrite in meat products by fluorescence quenching. For the sensing platform, carbon quantum dots doped with boron and functionalized with nitrogen (B,N-Cdot) were synthesized with an excellent 44.3% quantum yield via a one-step hydrothermal route using citric acid, boric acid, and branched polyethylenimine as carbon, boron, and nitrogen sources, respectively. After investigation of their chemical structure and fluorescent properties, the B,N-Cdot at aqueous suspensions showed high selectivity for NO2- in a linear range from 20 to 50 mmol L-1 under optimum conditions at pH 7.4 and a 340 nm excitation. Furthermore, the prepared B,N-Cdots successfully detected NO2- in a real meat sample with recovery of 91.4-104% within the analyzed range. In this manner, a B,N-Cdot/PVA nanocomposite film with blue emission under excitation at 360 nm was prepared, and a first assay detection of NO2- in meat products was tested using a smartphone application. The potential application of the newly developed sensing device containing a highly fluorescent probe should aid in the development of a rapid and inexpensive strategy for NO2- detection.