Visualization of Solvent Effect and Oxygen Content via a Red Room-Temperature Phosphorescent Material.

The development of pure organic room-temperature phosphorescent (RTP) materials greatly facilitates the integrated application of luminescent materials. Herein, a type of photoactivated red RTP material was constructed by simply doping 4-(benzo[c][1,2,5]thiadiazol-5-ylthio)benzonitrile (p-NNS) into a poly(methyl methacrylate) (PMMA) matrix. The obtained film realized a controllable photoactivation process by regulation of diverse solvent levels, demonstrating potential advantages in optical anti-counterfeiting applications. Furthermore, luminescent properties of the doped film were utilized to detect oxygen content from 2.00% to 4.90%, which revealed the exact consumption of ambient oxygen under UV light. Every CIE point of the luminescence corresponds to a certain oxygen content, illustrating the visualization of oxygen content. The remarkable regulation of solvent effect and oxygen content in this work will provide competitive material for further optical applications.

A portable NIR fluorimeter directly quantifies singlet oxygen generated by nanostructures for Photodynamic Therapy.

This paper reports on the setting up and calibration of a portable NIR fluorimeter specifically developed for quantitative direct detection of the highly reactive singlet oxygen (1O2) chemical specie, of great importance in Photodynamic therapies. This quantification relies on the measurement of fluorescence emission of 1O2, which is peaked in the near-infrared (NIR) at λ=1270nm. In recent years, several nanostructures capable of generating reactive oxygen species (ROS) when activated by penetrating radiation (X-rays, NIR light) have been developed to apply Photodynamic Therapy (PDT) to tumours in deep tissue, where visible light cannot penetrate. A bottleneck in the characterization of these nanostructures is the lack of a fast and reliable technique to quantitatively assess their performances in generating ROS, and in particular 1O2. For instance, the widely used PDT "Singlet Oxygen Sensor Green" kit suffers from self-activation under X-ray irradiation. To solve this difficulty, we propose here direct detection of 1O2 by spectroscopic means, using an apparatus developed by us around a recent thermoelectrically-cooled InGaAs single photon avalanche photodiode (SPAD). The SPAD is coupled to a custom-made integrating sphere designed for use under irradiation with high-energy X-ray beams from clinical Radiotherapy sources. We determine the detection threshold for our apparatus, which turns to be ∼9·1081O2 in realistic experimental condition and for measurements extending to 1 min of integration. After calibrations on standard photosensitizers, we demonstrate the potentiality of this instrument characterizing some photosensitizing nanostructures developed by us.

Fetal ischemia monitoring with in vivo implanted electrochemical multiparametric microsensors.

Under intrauterine growth restriction (IUGR), abnormal attainment of the nutrients and oxygen by the fetus restricts the normal evolution of the prenatal causing in many cases high morbidity being one of the top-ten causes of neonatal death. The current gold standards in hospitals to detect this relevant problem is the clinical observation by echography, cardiotocography and Doppler. These qualitative techniques are not conclusive and requires risky invasive fetal scalp blood testing and/or amniocentesis. We developed micro-implantable multiparametric electrochemical sensors for measuring ischemia in real time in fetal tissue and vascular. This implantable technology is designed to continuous monitoring for an early detection of ischemia to avoid potential fetal injury. Two miniaturized electrochemical sensors were developed based on oxygen and pH detection. The sensors were optimized in vitro under controlled concentration, to assess the selectivity and sensitivity required. The sensors were then validated in vivo in the ewe fetus model, by means of their insertion in the muscle leg and inside the iliac artery of the fetus. Ischemia was achieved by gradually obstructing the umbilical cord to regulate the amount of blood reaching the fetus. An important challenge in fetal monitoring is the detection of low levels of oxygen and pH changes under ischemic conditions, requiring high sensitivity sensors. Significant differences were observed in both; pH and pO2 sensors under changes from normoxia to hypoxia states in the fetus tissue and vascular with both sensors. Herein, we demonstrate the feasibility of the developed sensors for future fetal monitoring in medical applications.

Influence of Oxidative Stress on Time-Resolved Oxygen Detection by [Ru(Phen)3]2+ In Vivo and In Vitro.

Detection of tissue and cell oxygenation is of high importance in fundamental biological and in many medical applications, particularly for monitoring dysfunction in the early stages of cancer. Measurements of the luminescence lifetimes of molecular probes offer a very promising and non-invasive approach to estimate tissue and cell oxygenation in vivo and in vitro. We optimized the evaluation of oxygen detection in vivo by [Ru(Phen)3]2+ in the chicken embryo chorioallantoic membrane model. Its luminescence lifetimes measured in the CAM were analyzed through hierarchical clustering. The detection of the tissue oxygenation at the oxidative stress conditions is still challenging. We applied simultaneous time-resolved recording of the mitochondrial probe MitoTrackerTM OrangeCMTMRos fluorescence and [Ru(Phen)3]2+ phosphorescence imaging in the intact cell without affecting the sensitivities of these molecular probes. [Ru(Phen)3]2+ was demonstrated to be suitable for in vitro detection of oxygen under various stress factors that mimic oxidative stress: other molecular sensors, H2O2, and curcumin-mediated photodynamic therapy in glioma cancer cells. Low phototoxicities of the molecular probes were finally observed. Our study offers a high potential for the application and generalization of tissue oxygenation as an innovative approach based on the similarities between interdependent biological influences. It is particularly suitable for therapeutic approaches targeting metabolic alterations as well as oxygen, glucose, or lipid deprivation.

The effect of aging and antioxidants on photoreactivity and phototoxicity of human melanosomes: An in vitro study.

Aging may significantly modify antioxidant and photoprotective properties of melanin in retinal pigment epithelium (RPE). Here, photoreactivity of melanosomes (MS), isolated from younger and older human donors with and without added zeaxanthin and α-tocopherol, was analyzed by electron paramagnetic resonance oximetry, time-resolved singlet oxygen phosphorescence, and protein oxidation assay. The phototoxic potential of ingested melanosomes was examined in ARPE-19 cells exposed to blue light. Phagocytosis of FITC-labeled photoreceptor outer segments (POS) isolated from bovine retinas was determined by flow cytometry. Irradiation of cells fed MS induced significant inhibition of the specific phagocytosis with the effect being stronger for melanosomes from older than from younger human cohorts, and enrichment of the melanosomes with antioxidants reduced the inhibitory effect. Cellular protein photooxidation was more pronounced in samples containing older melanosomes, and it was diminished by antioxidants. This study suggests that blue light irradiated RPE melanosomes could induce substantial inhibition of the key function of the cells-their specific phagocytosis. The data indicate that while photoreactivity of MS and their phototoxic potential increase with age, they could be reduced by selected natural antioxidants.

Oxygen Gas Sensing with Photothermal Spectroscopy in a Hollow-Core Negative Curvature Fiber.

We demonstrate a compact all-fiber oxygen sensor using photothermal interferometry with a short length (4.3 cm) of hollow-core negative curvature fibers. The hollow-core fiber has double transmission windows covering both visible and near-infrared wavelength regions. Absorption of a pump laser beam at 760 nm produces photothermal phase modulation and a probe Fabry-Perot interferometer operating at 1550 nm is used to detect the phase modulation. With wavelength modulation and first harmonic detection, a limit of detection down to 54 parts per million (ppm) with a 600-s averaging time is achieved, corresponding to a normalized equivalent absorption of 7.7 × 10-8 cm-1. The oxygen sensor has great potential for in situ detection applications.

Fabrication, Characterization, and Analytical Application of Silica Nanopore Array-Modified Platinum Electrode.

In this work, we report a new approach to fabricate the nanopore array electrode (NAE) by transferring silica nanochannel membrane (SNM) to the surface of Pt electrode (0.5 mm in diameter) sealed by glass capillary (designated as Pt-NAE for simplicity). The SNM is supported via the irreversible covalent-bond formation with the surrounding glass capillary treated by plasma, thus providing long-term stability to Pt-NAE. Meanwhile, this fabrication process does not require pregrafting or premodification of Pt electrode surface, providing well-defined active surface domains. Thanks to the small pore diameter (∼2.3 nm) and negatively charged channel walls, the SNM is permselective and thus the electrochemical behavior of Pt-NAE is dependent on both electrolyte concentration and charge state of redox molecules. The permeability of SNM was determined by the scanning electrochemical microscopy (SECM) approach curve measurements coupled with finite-element simulations from a quantitative viewpoint. The permeability of anionic Ru(CN)64- was varied from 150 to 10.3 µm s-1 as the electrolyte concentration decreased from 1.0 to 0.01 M, while there is no obvious change for cationic Ru(NH3)63+. Finally, the as-prepared Pt-NAE is able to continuously monitor dissolved oxygen for up to 2 h in a solution containing biofouling reagents, exhibiting an enhanced antifouling ability and therefore excellent current stability. We believe the NAE with unique mass transport properties can be extended further for other analytical applications.

Oxygen-Sensitive Layered MoTe2 Channels for Environmental Detection.

The oxygen (O2)-dependent resistance change of multilayered molybdenum ditelluride (MoTe2) channels was characterized. A variation of the channel resistance could reproducibly determine relative O2 content (denoted as the O2 index). We found that Joule heating in a layered MoTe2 field-effect transistor caused the O2 index to decrease drastically from 100 to 12.1% in back gate modulation. Furthermore, Joule heating caused effective O2 desorption from the MoTe2 surface and repeatable O2 detection by multilayered MoTe2 channels was realized. This work not only explored the influence of O2 on the electrical properties of multilayered MoTe2 channels but also revealed that MoTe2 channels are promising for sensing O2 in an environmental condition.

Polylysine modified conjugated polymer nanoparticles loaded with the singlet oxygen probe 1,3-diphenylisobenzofuran and the photosensitizer indocyanine green for use in fluorometric sensing and in photodynamic therapy.

Conjugated polymer hybrid nanoparticles (NPs) loaded with both indocyanine green (ICG) and 1,3-diphenylisobenzofuran (DPBF) are described. The NPs are dually functional in that ICG acts as the photosensitizer, and DPBF as a probe for singlet oxygen (1O2 probe). The nanoparticle core consists of the energy donating host poly(9,9-dioctylfluorenyl-2,7-diyl)-co-(2,5-p-xylene) (PFP). The polymer is doped with the energy acceptor DPBF. Ratiometric fluorometric detection of singlet oxygen is accomplished by measurement of fluorescence at wavelengths of 415 and 458 nm. In addition, the shell of the positively charged polymeric nanoparticles was modified, via electrostatic interaction, with negatively charged PDT drugs ICG. The integrated nanoparticles of type ICG-DPBF-PFP display effective photodynamic performance under 808-nm laser irradiation. The 1O2 sensing behaviors of samples are evaluated based on the ratiometric fluorescent responses produced by DPBF and PFP. 1O2 can be fluorimetically sensed with a detection limit of 28 µM. The multifunctional nanoprobes exhibit effortless cellular uptake, superior photodynamic activity and a rapid ratiometric response to 1O2. Graphical abstractSchematic of a dual-functional nanoplatform for photodynamic therapy (PDT) and singlet oxygen (1O2) feedback. It offers a new strategy for self-monitoring photodynamic ablation. FRET: fluorescence resonance energy transfer. Indocyanine green is attached in the shell of nanoparticles, and 1,3-diphenylisobenzofuran is doped into the energy donating host conjugated polymer.

Enhanced sensitivity of scanning bipolar electrochemical microscopy for O2 detection.

The Scanning Bipolar Electrochemical Microscope (SBECM) allows precise positioning of an electrochemical micro-probe serving as bipolar electrode that can be wirelessly interrogated by coupling the electrochemical detection reaction with an electrochemiluminescent reporting process. As a result, the spatially heterogeneous concentrations of an analyte of interest can be converted in real time into a map of sample reactivity. However, this can only be achieved upon optimization of the analytical performance ensuring adequate sensitivity. Here, we present the evaluation and optimized operation of the SBECM for the detection of small changes in local O2 concentrations. Parameters for achieving an improved sensitivity as well as possibilities for improving the signal-to-noise ratio in the optical signal readout are evaluated. The capability of the SBECM for O2 detection is shown at controlled conditions by recording the topography of a patterned sample and monitoring O2 evolution from a photoelectrocatalyst material.

Long-Lived Room-Temperature Phosphorescence for Visual and Quantitative Detection of Oxygen.

An unconventional organic molecule (TBBU) showing obvious long-lived room temperature phosphorescence (RTP) is reported. X-ray single crystal analysis demonstrates that TBBU molecules are packed in a unique fashion with side-by-side arranged intermolecular aromatic rings, which is entirely different from the RTP molecules reported to date. Theoretical calculations verify that the extraordinary intermolecular interaction between neighboring molecules plays an important role in RTP of TBBU crystals. More importantly, the polymer film doped with TBBU inherits its distinctive RTP property, which is highly sensitive to oxygen. The color of the doped film changes and its RTP lifetime drops abruptly through a dynamic collisional quenching mechanism with increasing oxygen fraction, enabling visual and quantitative detection of oxygen. Through analyzing the grayscale of the phosphorescence images, a facile method is developed for rapid, visual, and quantitative detection of oxygen in the air.

Sensitive, Real-Time, and In-Vivo Oxygen Monitoring for Photodynamic Therapy by Multifunctional Mesoporous Nanosensors.

Real-time monitoring of oxygen consumption is beneficial to predict treatment responses and optimize therapeutic protocols for photodynamic therapy (PDT). In this work, we first demonstrate that deformable hollow mesoporous organosilica nanoparticles (HMONs) can be used to load [(Ru(dpp)3)]Cl2 for detecting oxygen (denoted as HMON-[(Ru(dpp)3)]Cl2). This nanoprobe shows significantly improved biocompatibility and high cellular uptake. In-vitro experiments demonstrate that the HMON-[(Ru(dpp)3)]Cl2 can sensitively detect oxygen changes between 1% and 20%. On this basis, photosensitizer chlorin e6 (Ce6) and [(Ru(dpp)3)]Cl2 are simultaneously loaded in the HMONs (denoted as HMON-Ce6-[(Ru(dpp)3)]Cl2) for real-time oxygen monitoring during photodynamic therapy. The HMON-Ce6-[(Ru(dpp)3)]Cl2 can reflects oxygen consumption in solution and cells in photodynamic therapy. Furthermore, the ability of the HMON-Ce6-[(Ru(dpp)3)]Cl2 nanosensor to monitor oxygen changes is demonstrated in tumor-bearing nude mice.

Hypoxic Physiological Environments in a Gas-Regulated Microfluidic Device.

Hypoxic environment is known as one of the critical factors in various physiological/pathological processes. It is imperative to recapitulate oxygen level in microscale for human physiology/pathology induced by hypoxia. Herein, we propose an oxygen-regulating system that can be applied to in vitro tissue models. We fabricated a microdevice with a gas-permeable membrane, allowing oxygen diffusion without direct contact to cells. We verified the formation of oxygen level less than 2% O2 concentration inside the device through computational simulation and experiments. H9c2 heart myoblasts were exposed to hypoxic condition in the device, and their cell viability were investigated. We anticipate that our system will be integrated with a platform to study hypoxia-induced human physiology and pathology as an efficient oxygen-regulating system.

Redox-Triggered Bonding-Induced Emission of Thiol-Functionalized Gold Nanoclusters for Luminescence Turn-On Detection of Molecular Oxygen.

Most optical sensors for molecular oxygen were developed based on the quenching effect of the luminescence of oxygen-sensitive probes; however, the signal turn-off mode of these probes is undesirable to quantify and visualize molecular oxygen. Herein, we report a novel luminescence turn-on detection strategy for molecular oxygen via the specific oxygen-triggered bonding-induced emission of thiol-functionalized gold nanoclusters. Thiol-functionalized gold nanoclusters were prepared by a facile one-step synthesis, and as-prepared gold nanoclusters possess significant aggregation-induced emission (AIE) property. It is the first time to discover the oxygen-triggered bonding-induced emission (BIE) behavior of gold nanoclusters, which results in disulfide-linked covalent bonding assemblies with intensely red luminescence. This specific redox-triggered BIE is capable of quantitatively detecting dissolved oxygen in aqueous solution in a light-up manner, and trace amount of dissolved oxygen at ppb level is achieved based on this detection method. A facile and convenient test strip for oxygen detection was also developed to monitor molecular oxygen in a gas matrix. Covalent bonding-induced emission is proven to be a more efficient way to attain high brightness of AIEgens than a physical aggregation-induced emission process, and provides a more convenient and desirable detection method for molecular oxygen than the previous sensors.

Lipofuscin-mediated photic stress inhibits phagocytic activity of ARPE-19 cells; effect of donors' age and antioxidants.

The risk of chronic oxidative stress in the retinal pigment epithelium (RPE) increases with age due to accumulation of the photoreactive age pigment lipofuscin (LFG). Here, we asked whether sublethal and weakly lethal photic stress, induced by irradiation of ARPE-19 cells containing phagocytised LFG, affected the cell specific phagocytic activity, which is critically important for proper functioning and survival of the retina, and if natural antioxidants could modify the observed outcomes. ARPE-19 cells preloaded with LFG isolated from human donors of different age or containing LFG enriched with zeaxanthin and α-tocopherol (LFG-A), were irradiated with blue light. Phagocytosis of fluorescein-5-isothiocyanate (FITC)-labelled photoreceptor outer segments was determined by flow cytometry. Photoreactivity of LFG and LFG-A was analysed by measuring photoconsumption of oxygen and photogeneration of singlet oxygen mediated by the granules. LFG-mediated photic stress in ARPE-19 cells induced significant inhibition of their specific phagocytosis. The inhibitory effect increased with age of LFG donors and was reduced by enrichment of the granules with antioxidants. Oxygen consumption and generation of singlet oxygen induced by the photoexcited LFG increased with donor's age and was partially quenched by antioxidants. Although the phototoxic potential of lipofuscin increased with age, natural antioxidants reduced photoreactivity of LFG and their efficiency to induce oxidative stress. This study has demonstrated, for the first time, that mild oxidative stress, mediated by the age pigment lipofuscin, impairs specific phagocytic activity of RPE, and that natural antioxidants can protect this important cellular function by reducing lipofuscin photoreactivity.

Rapid Measurement of Room Temperature Ionic Liquid Electrochemical Gas Sensor using Transient Double Potential Amperometry.

Intense study on gas sensors has been conducted to implement fast gas sensing with high sensitivity, reliability and long lifetime. This paper presents a rapid amperometric method for gas sensing based on a room temperature ionic liquid electrochemical gas sensor. To implement a miniaturized sensor with a fast response time, a three electrode system with gold interdigitated electrodes was fabricated by photolithography on a porous polytetrafluoroethylene substrate that greatly enhances gas diffusion. Furthermore, based on the reversible reaction of oxygen, a new transient double potential amperometry (DPA) was explored for electrochemical analysis to decrease the measurement time and reverse reaction by-products that could cause current drift. Parameters in transient DPA including oxidation potential, oxidation period, reduction period and sample point were investigated to study their influence on the performance of the sensor. Oxygen measurement could be accomplished in 4 s, and the sensor presented a sensitivity of 0.2863 µA/[%O2] and a linearity of 0.9943 when tested in air samples with different oxygen concentrations. Repeatability and long-term stability were also investigated, and the sensor was shown to exhibit good reliability. In comparison to conventional constant potential amperometry, transient DPA was shown to reduce relative standard deviation by 63.2%. With transient DPA, the sensitivity, linearity, repeatability, measurement time and current drift characteristics demonstrated by the presented gas sensor are promising for acute exposure applications.

Different valence Sn doping - A simple way to detect oxygen concentration variation of ZnO quantum dots synthesized under ultrasonic irradiation.

An ultrasonic method is employed to synthesize the Sn doped Zn0.95Sn0.05O quantum dots with green light emission. Sn2+ and Sn4+ ions are used to create different optical defects inside Zn0.95Sn0.05O quantum dots and the changing trend of oxygen concentration under different ultrasonic irradiation power are investigated. The photoluminescence spectra are employed to characterize the optical defects of Zn0.95Sn0.05O quantum dots. The UV-vis spectra are used to study the band gap of Zn0.95Sn0.05O quantum dots, which is influenced by their sizes. The results indicate that ultrasonic power would influence the size of Zn0.95Sn0.05O quantum dots as well as the type and quantity of defects in ZnO quantum dots. Changing trends in size of Sn2+ and Sn4+ doped Zn0.95Sn0.05O quantum dots are quite similar with each other, while the changing trends in optical defects types and concentration of Sn2+ and Sn4+ doped Zn0.95Sn0.05O quantum dots are different. The difference of the optical defects concentration changing between Sn2+ doped Zn0.95Sn0.05O quantum dots (VO defects) and Sn4+ doped Zn0.95Sn0.05O quantum dots (OZn and Oi defects) shows that the formation process of ZnO under ultrasonic irradiation wiped oxygen out.

Targeting tumour hypoxia to prevent cancer metastasis. From biology, biosensing and technology to drug development: the METOXIA consortium.

The hypoxic areas of solid cancers represent a negative prognostic factor irrespective of which treatment modality is chosen for the patient. Still, after almost 80 years of focus on the problems created by hypoxia in solid tumours, we still largely lack methods to deal efficiently with these treatment-resistant cells. The consequences of this lack may be serious for many patients: Not only is there a negative correlation between the hypoxic fraction in tumours and the outcome of radiotherapy as well as many types of chemotherapy, a correlation has been shown between the hypoxic fraction in tumours and cancer metastasis. Thus, on a fundamental basis the great variety of problems related to hypoxia in cancer treatment has to do with the broad range of functions oxygen (and lack of oxygen) have in cells and tissues. Therefore, activation-deactivation of oxygen-regulated cascades related to metabolism or external signalling are important areas for the identification of mechanisms as potential targets for hypoxia-specific treatment. Also the chemistry related to reactive oxygen radicals (ROS) and the biological handling of ROS are part of the problem complex. The problem is further complicated by the great variety in oxygen concentrations found in tissues. For tumour hypoxia to be used as a marker for individualisation of treatment there is a need for non-invasive methods to measure oxygen routinely in patient tumours. A large-scale collaborative EU-financed project 2009-2014 denoted METOXIA has studied all the mentioned aspects of hypoxia with the aim of selecting potential targets for new hypoxia-specific therapy and develop the first stage of tests for this therapy. A new non-invasive PET-imaging method based on the 2-nitroimidazole [(18)F]-HX4 was found to be promising in a clinical trial on NSCLC patients. New preclinical models for testing of the metastatic potential of cells were developed, both in vitro (2D as well as 3D models) and in mice (orthotopic grafting). Low density quantitative real-time polymerase chain reaction (qPCR)-based assays were developed measuring multiple hypoxia-responsive markers in parallel to identify tumour hypoxia-related patterns of gene expression. As possible targets for new therapy two main regulatory cascades were prioritised: The hypoxia-inducible-factor (HIF)-regulated cascades operating at moderate to weak hypoxia (<1% O(2)), and the unfolded protein response (UPR) activated by endoplasmatic reticulum (ER) stress and operating at more severe hypoxia (<0.2%). The prioritised targets were the HIF-regulated proteins carbonic anhydrase IX (CAIX), the lactate transporter MCT4 and the PERK/eIF2α/ATF4-arm of the UPR. The METOXIA project has developed patented compounds targeting CAIX with a preclinical documented effect. Since hypoxia-specific treatments alone are not curative they will have to be combined with traditional anti-cancer therapy to eradicate the aerobic cancer cell population as well.