Fluorescence Lifetime Analysis Research Articles

Optimizing optical thermometry with tri-doped Ba2GdV3O11 phosphors: Ratiometric and fluorescence lifetime analysis

Optical sensor technology has undergone a transformative evolution with the advent of fluorescence ratio techniques (FIR) and fluorescence lifetime (FL) strategies, revolutionizing precision, performance, and reliability. This study delves into the synthesis of Ba2GdV3O11 phosphors doped with Ho3+/Nd3+, Er3+, and Yb3+, employing the sol-gel method for upconverting material fabrication. A thorough investigation into the structural, morphological, and optical properties of the synthesized phosphors is conducted. Excitation at 980 nm unveils upconversion (UC) emissions across green and red spectra. The intensities of the observed emission bands for Ho3+, Nd3+, and Er3+ demonstrate significant sensitivity to fluctuations in temperature. Temperature sensing relies on the 4S3/2 and 2H11/2 upconversion emissions bands, in addition to the emission lifetimes at 4S3/2. Enhanced thermal sensitivity values are attained, reaching up to 1.03 % K−1 and 1.07 % K−1 using the FIR strategy, and up to 0.146 % K−1 and 0.47 % K−1 with the FL strategy for Ho3+/Er3+/Yb3+ and Nd3+/Er3+/Yb3+ tri-doped Ba2GdV3O11 phosphors, respectively. Furthermore, the studied phosphors exhibit remarkable precision in detecting minute temperature changes (0.3 K), positioning them as promising candidates for precise temperature sensing. This study pioneers innovative methodologies to advance optical thermometry techniques, offering promising prospects for scientific and industrial applications reliant on precise optical temperature sensing.

A time-correlated single photon counting SPAD array camera with a bespoke data-processing algorithm for lightsheet fluorescence lifetime imaging (FLIM) and FLIM videos

A wide-field microscope with epi-fluorescence and selective plane illumination was combined with a single-photon avalanche diode (SPAD) array camera to enable live-cell fluorescence lifetime imaging (FLIM) using time-correlated single-photon counting (TCSPC). The camera sensor comprised of 192×128\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\ ext {192}\ imes \ ext {128}$$\\end{document} pixels, each integrating a single SPAD and a time-to-digital converter. Jointly, they produced a stream of single-photon images of photon arrival times with ≈38 ps\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\approx \ ext {38 ps}$$\\end{document} accuracy. The photon arrival times were subject to systematic delays and nonlinearities, which were corrected by a Monte-Carlo algorithm. The SPAD camera was then applied to FLIM where histogramming the resulting photon arrival times in each pixel resulted in decays compatible with common data processing pipelines for fluorescence lifetime analysis. The capabilities of the TCSPC camera-based FLIM microscope were demonstrated by imaging living unicellular photosynthetic algae and artificial lipid vesicles. Epi-fluorescence illumination enabled rapid fluorescence lifetime imaging of living cells and selective-plane illumination enabled 3-dimensional FLIM of stationary samples.

A new yellowish‐green‐emitting phosphor: Eu2+‐doped Ba3YAl2O7.5

AbstractA yellowish‐green emitting phosphor Ba3YAl2O7.5:Eu2+ was synthesized by a solid‐state method. The crystal structure, luminescence, concentration quenching, fluorescence decay, and thermal quenching properties were systematically investigated. Under blue light (440 nm) excitation, Ba3YAl2O7.5:Eu2+ phosphors exhibit a yellowish‐green emission peaking at 550 nm with a full width at half maximum ∼ 69 nm, and The Commission Internationale de l'Eclairage (CIE) coordinates of Ba3YAl2O7.5:Eu2+ are (0.4064, 0.5811). Structural analysis and Van Uitert equation along with fluorescence lifetime analysis reveal that Eu2+ ions occupy 6‐coordinated Y and 9‐coordinated Ba sites in Ba3YAl2O7.5, and therefore generate two emission sub‐bands at 582 and 548 nm. In addition, Ba3YAl2O7.5:Eu2+ phosphor remaining 43% photoluminescence emission intensity at 125°C with an activation energy of 0.372 eV. Utilizing the title phosphor (Ba3YAl2O7.5:3%Eu2+), commercial red (Ca,Sr)AlSiN3:Eu2+ (CSAlSi:Eu2+) phosphor, and InGaN‐based blue light emitting diode (LED) chip, a proof‐of‐concept warm white LEDs (WLEDs) with a color rendering index of 80.1, CIE chromaticity coordinates of (0.411, 0.381), luminous efficacy of ∼ 22.13 lm/W, and correlated color temperature of ∼ 3288 K is achieved. Thus, Eu2+ activated Ba3YAl2O7.5 phosphors have the potential application in WLEDs pumped by a blue LED chip for solid‐state lighting application.

Visible light assisted photocatalytic performance of Europium doped ZnS-Nb2O5 heterostructure for mineralization of Acid violet 7 dye

The present study focuses on the visible light assisted photocatalytic performance of Europium doped ZnS-Nb2O5 heterostructure catalysts for the mineralization of organic pollutants such as Acid violet 7 (AV7) dye. Structural and morphological investigations reveal the presence of wurtzite ZnS and Nb2O5 components in the prepared heterostructure in separate phases. The prepared Europium doped ZnS-Nb2O5 (3Eu50ZN) heterostructures shows enhanced visible light absorption with prominent redshift. Photoluminescence study suggests the presence of surface states/defect energy levels in Europium doped ZnS-Nb2O5 heterostructure catalysts. In comparison to doped and undoped components, the fluorescence lifetime analysis demonstrates enhanced charge separation in 3Eu50ZN heterostructure catalysts. Photo/electrochemical studies reveal superior capacitance behavior, enhanced charge carrier separation, and improved stability in the aqueous phase for 3Eu50ZN heterostructure in comparison to ZnS and Nb2O5 catalysts. The superior visible light assisted photocatalytic performance with 3Eu50ZN heterostructure catalysts was achieved for the mineralization of AV7 dye (20 mg L−1). Active species determination suggests photogenerated holes (h+) to be the major active species responsible for the degradation reaction and a plausible reaction pathway was proposed. The 3Eu50ZN heterostructure catalysts exhibit a recycling nature and demonstrate improved stability.

The effect of membrane composition on the interaction between human CYP51 and its flavonoid inhibitor - luteolin 7,3′-disulfate

Cytochromes P450 (CYP) are a family of membrane proteins involved in the production of endogenous molecules and the metabolism of xenobiotics. It is well-known that the composition of the membrane can influence the activity and orientation of CYP proteins. However, little is known about how membrane composition affects the ligand binding properties of CYP. In this study, we utilized surface plasmon resonance and fluorescence lifetime analysis to examine the impact of membrane micro-environment composition on the interaction between human microsomal CYP51 (CYP51A1) and its inhibitor, luteolin 7,3′-disulphate (LDS). We observed that membranes containing cholesterol or sphingomyelin exhibited the lowest apparent equilibrium dissociation constant for the CYP51A1-LDS complex. Additionally, the tendency for relation between kinetic parameters of the CYP51A1-LDS complex and membrane viscosity and overall charge was observed. These findings suggest that the specific composition of the membrane, particularly the presence of cholesterol and sphingomyelin, plays a vital role in regulating the interaction between CYP enzymes and their ligands.

Fluorescence Lifetime Measurements and Analyses: Protocols Using Flow Cytometry and High-Throughput Microscopy.

This chapter focuses on applications and protocols that involve the measurement of the fluorescence lifetime as an informative cytometric parameter. The timing of fluorescence decay has been well-studied for cell counting, sorting, and imaging. Therefore, provided herein is an overview of the techniques used, how they enhance cytometry protocols, and the modern techniques used for lifetime analysis. The background and theory behind fluorescence decay kinetic measurements in cells is first discussed followed by the history of the development of time-resolved flow cytometry. These sections are followed by a review of applications that benefit from the quantitative nature of fluorescence lifetimes as a photophysical trait. Lastly, perspectives on the modern ways in which the fluorescence lifetime is scanned at high throughputs which include high-speed microscopy and machine learning are provided.

Fast and portable fluorescence lifetime analysis for early warning detection of micro- and nanoplastics in water

The presence of plastic fragments in aquatic environments, particularly at the micro- and nano-scale, has become a significant global concern. However, current detection methods are limited in their ability to reveal the presence of such particles in liquid samples. In this study, we propose the use of a fluorescence lifetime analysis system for the detection of micro- and nanoplastics in water. This approach relies on the inherent endogenous fluorescence of plastic materials and involves the collection of single photons emitted by plastic fragments upon exposure to a pulsed laser beam. Briefly, a pulsed laser beam (repetition frequency = 40 MHz) shines onto a sample solution, and the emitted light is filtered, collected, and used to trace the time distributions of the photons with high temporal resolution. Finally, the fluorescence lifetime was measured using fitting procedures and a phasor analysis. Phasor analysis is a fit-free method that allows the measurement of the fluorescence lifetime of a sample without any assumptions or prior knowledge of the sample decay pattern. The developed instrument was tested using fluorescence references and validated using unlabelled micro- and nano-scale particles. Our system successfully detected polystyrene particles in water, achieving a remarkable sensitivity with a detection limit of 0.01 mg/mL, without the need for sample pre-treatment or visual inspection. Although further studies are necessary to enhance the detection limit of the technique and distinguish between different plastic materials, this proof-of-concept study suggests the potential of the fluorescence lifetime-based approach as a rapid, robust, and cost-effective method for early warning detection and identification of plastic contaminants in aquatic environments.

Highly sensitive and selective off-on fluorescent platform for tricresyl phosphate flame retardant based on twisted intramolecular charge transfer probe

BackgroundTricresyl phosphate (TCP), a typical organic phosphorus flame retardant (OPFR), is an emerging pollutant that causes great concern in recent years due to its high neurotoxicity and reproductive toxicity, etc. Conventional analysis methods for TCP such as gas chromatography and liquid chromatography-tandem mass spectrometry exhibit high sensitivity and accuracy. However, these techniques generally suffer from certain limitations, such as high cost, bulky equipment, time-consuming and operator-dependent properties. Therefore, the establishment of fast and efficient analytical methods for TCP still remains a great challenge. ResultA “turn on” fluorescence sensing strategy for the efficient detection of TCP was established, based on a unique molecular rotor probe of 9-(2,2-dicyanovinyl)-julolidine (DCVJ). The introduction of TCP led to a significant enhancement of the fluorescence intensity of DCVJ. The results show that twisted intramolecular charge transfer (TICT) process might play an important role for this enhancement of fluorescence response via dynamic light scattering measurements and fluorescence lifetime analysis. Further investigations demonstrate that the hydrophobic interaction and conjugation effect between DCVJ and TCP constrain the molecular rotation and vibration of DCVJ, thereby regulating the TICT process, which contribute to this intriguing “turn on” behavior. In view of this, a new sensing platform with excellent performance for TCP was established, which offers quick response time, high selectivity, wide linear range (20–1200 ng mL−1, 1600–8000 ng mL−1), and low detection limit (4.82 ng mL−1). SignificanceThe established new sensing platform for TCP demonstrates the advantages of simplicity, high efficiency, excellent sensitivity and selectivity. The obtained results are also superior to some other previously reported fluorescence methods. This work opens up a new perspective for the efficient detection of emerging OPFRs pollutants.

Exploration of the crystal growth, spectroscopy, and LD-pumped 2.7 μm laser operation of Er:LuScO3 sesquioxide crystals

High quality sesquioxide Er:LuScO3 crystals were grown successfully by the laser floating zone method. Based on the analysis of luminescence spectra and fluorescence lifetimes at the temperatures of 77 K and 300 K. The Er3+-doped sesquioxide shows an effective spectral broadening in the solid solution mixing. Additionally, the local polarization and electron-phonon coupling will be impacted by the increase of Er3+ ions, both of which are favorable for the broadening of spectra. The 10 at% Er:LuScO3 was optimized to operate the 2.7 μm laser, and the largest output power of 326 mW can be acquired with a slope efficiency of 11.3%. To the best of our knowledge, this work was the first to demonstrate a diode-end-pumped Er:LuScO3 crystal laser operation around 3.0 μm.

Crystal growth, HOMO-LUMO engineering and charge-transfer emission of TCNB-based donor-acceptor cocrystals

Using carbazole (CARB), dibenzothiophene (DBZP) and dibenzofuran (DBZF) as electron donors, 1,2,4,5-tetracyanobenzene (TCNB) as electron acceptor, three typical types of luminescent organic cocrystals were synthesized by a simple supramolecular assembly strategy. The cocrystals exhibit interesting optical properties for its charge transfer (CT) interactions. Meanwhile, a series of three-component (CARB)x (DBZP)1-x (TCNB) crystalline microtubes comprising organic CT interactions have been prepared by the self-assembly with poor solvent co-precipitating method. DFT calculations were used to predict the CT interactions and the binding patterns of donor and acceptor, which suggest that the CT interactions can largely influence the energy level and orbital distributions. Moreover, the SXRD analyses verified the cocrystals are constructed by π-π stacking with a face-to-face model. The solid-state cocrystals and microtubes were well characterized by PXRD, UV absorption spectra, emission spectra, fluorescence lifetime and quantum yield analyses. The obvious red-shift in UV–vis absorption spectra and emission spectra with respect to the free monomer indicate the formation of CT interactions. The emission colors of the mixed microtubes can be tailored from green to orange as well as associated photophysical properties with the increasing of dopant concentrations. Therefore, by the combination of experimental and theoretical studies on the cocrystals and microtubes, this work not only reports the new types of crystalline micro-nano materials, but also provides an in-depth understanding of the relationship between structure and fluorescence effect.

A water-stabilized Tb-MOF can be used as a sensitive and selective fluorescence sensor for the detection of oxytetracycline hydrochloride

Oxytetracycline hydrochloride (OTC) is a commonly used over-the-counter antibiotic, valued for its potent antibacterial properties. However, the inappropriate and excessive use of OTC can result in the accumulation of the drug in both the environment and human body, causing significant harm to ecosystems and human health. Therefore, the development of a fast and sensitive method for detecting OTC is of great significance. Lanthanide metal–organic frameworks (LnMOFs) can effectively excite lanthanide metals to emit long-lifetime, narrow and stable fluorescence based on the antenna effect, but their application in fluorescence sensing is rarely reported. In this work, a strongly fluorescent material Tb-MOF was synthesized by a facile solvothermal method using 1,2,4,5-phenylenetetracarboxylic acid (H4btec) and 1,10-phenanthroline (1,10-phen) as organic ligands and lanthanide metal Tb as the luminescent center. Tb-MOF is a stable material in water and shows excellent linearity with OTC in the concentration range of 0 ∼ 70 μM, with low detection limit (0.12 μM) and luminescence color transition from bright green to dark green during the detection process. X-ray diffraction, UV–vis absorption and fluorescence lifetime analyses revealed that the fluorescence quenching of Tb-MOF by OTC is caused by the inner filter effect in static quenching. Test strips for OTC detection were successfully prepared using Tb-MOF. These strips are not only low cost and easy to prepare but can also be used as portable sensing devices that can be easily distinguished by the naked eye during OTC testing. This study not only presents a fluorescent probe for the detection of OTC in water but also offers a practical method for converting fluorescent luminescent materials into functional devices for OTC detection.

Two-site occupancy induced a broadband blue-emitting in Eu2+ doped Na3Ba2Ca(PO4)3 phosphor

During the process of developing commercial chips, the phosphors with single broadband emission are indispensable. In this report, a new ternary phosphate phosphor Na3Ba2Ca(PO4)3:Eu2+ has been successfully synthesized by the high-temperature solid-state reaction (HSSR) method, which shows broad emitting band in the range of 350–600 nm centered at 436 nm (FWHM ∼ 72.3 nm). The crystal structure of the Na3Ba2Ca(PO4)3 and Eu2+ doped Na3Ba2Ca(PO4)3 were determined by the Rietveld method, indicating the trigonal system with P31c space group. The selective two-site occupancy tendency of Eu2+ ions at Ba1O9 and Ca2O8 sites in the matrix was comprehensively revealed according to the split peaks (433 nm and 469 nm) at 298 K and 11 K, computational validation and fluorescence lifetime analysis of photoluminescence spectrum. Impressively, the optimal Na3Ba2Ca(PO4)3:0.02Eu2+ phosphor exhibits appropriate spectrum resemblance (SR) indexes (58.5% and 72.2%), proper thermostability (78.6%) and excellent chromaticity stability (Δx = 8.68 × 10−6 and Δy = 3.64 × 10−5). Besides, the phosphor prepared by HSSR owns the higher internal quantum efficiency of 62.1% in comparison with that synthesized by polymerizable complex (PC) method. The new phosphor can serve as a potential broadband and blue-emitting phosphor for the application of solid-state lighting and plant growth.

Achieving chip-matched high purity red emission by Bi3+ sensitization in Ca3MgTiGe3O12:Pr3+

Phosphor-converted light emitting diodes (pc-LEDs) are essential for current lighting and display areas because of their unique properties of being uncomplicated, durable and economical. In previous studies, Pr3+ doped red-emitting phosphors have attracted enormous research interest due to their high color purity. However, their photoluminescence excitation (PLE) spectra are always hard to be combined with LED chips. This mismatch is a great obstacle to achieve Pr3+ phosphors utilizing in LED devices. Here, to circumvent this issue, Pr3+-doped Ca3MgTiGe3O12 (CMTG) red-emitting phosphor was optimized with PLE band shifting from 330 nm to 360 nm after tuning with Bi3+ co-doping based on the structure and phosphorescence properties. The chip-matched excitation makes it a good candidate for white LEDs (wLEDs) fabrication to improve the output quality of device. Meanwhile, the high color purity of 93.02 % was obtained, which is conducive to widening the color gamut of backlight display applications. The photoluminescence (PL) spectra, fluorescence lifetime, and X-ray photoelectron spectroscopy (XPS) analysis were employed and discussed in detail to explain the role of Bi3+ played in the system. Finally, wLEDs were fabricated with CMTG:Pr3+, Bi3+, showing a high color rendering index (Ra) of 94.4. And the color gamut covers ∼80 % of the National Television System Committee (NTSC) standard, suggesting great potential for applications of wLEDs and backlight display.

Optical and DFT study of a novel blue-emitting Gd7O6(BO3) (PO4)2: Bi3+ phosphor

In this study, a novel blue-emitting phosphors, Gd7O6(BO3) (PO4)2: Bi3+, has been successfully synthesized via a high-temperature solid-state method, and first-principles calculations, microscopic structures, and optical properties of these phosphors were studied. The bandgap and density of states in both the undoped and doped states by implementing the Vienna Ab initio Simulation Package were also investigated. The obtained results indicated that the introduction of Bi3+ dopants led to a reduction in the bandgap value, as ascertained through data analysis. Based on the spectroscopic analysis, it can be inferred that the material A manifests a Stoke shift of 10,073 cm−1 and full width at half maximum (FWHM) of 87 nm. The thermal stability of the spectrum indicates a slight blue shift with increasing temperature, accompanied by a small increase in FWHM. Based on the fluorescence lifetime analysis, it has been revealed that there is a reduction in the fluorescence lifetime with the escalation of the doping concentration. The CIE diagram displays that all the samples at different concentrations are placed in the blue region and primarily converge at a single point.

Formation of a stable PSI-PSII megacomplex in rice that conducts energy spillover.

In green plants, photosystem I (PSI) and photosystem II (PSII) bind to their respective light-harvesting complexes (LHCI and LHCII) to form the PSI-LHCI supercomplex and PSII-LHCII supercomplex, respectively. These supercomplexes further form megacomplexes, like the PSI-PSII and PSII-PSII in Arabidopsis (Arabidopsis thaliana) and spinach to modulate their light-harvesting properties, but not in the green alga Chlamydomonas reinhardtii. Here, we fractionated and characterized the stable rice PSI-PSII megacomplex. The delayed fluorescence from PSI (lifetime ~ 25 ns) indicated energy transfer capabilities between the two photosystems (energy spillover) in the rice PSI-PSII megacomplex. Fluorescence lifetime analysis revealed that the slow PSII to PSI energy transfer component was more dominant in the rice PSI-PSII supercomplexes than in Arabidopsis ones, suggesting that the PSI and PSII in rice form a megacomplex not directly but through LHCII molecule(s), which was further confirmed by negatively-stained EM-analysis. Our results suggest species diversity in the formation and stability of photosystem megacomplexes, and the stable PSI-PSII supercomplex in rice may reflect its structural adaptation.

A Novel and Sensitive Fluorescent Probe for Glyphosate Detection Based on Cu2+ Modulated Polydihydroxyphenylalanine Nanoparticles.

A novel and sensitive fluorescent probe based on Cu2+-modulated polydihydroxyphenylalanine nanoparticles (PDOAs) has been developed for the detection of glyphosate pesticides. Compared to conventional instrumental analysis techniques, fluorometric methods have obtained good results in the field of agricultural residue detection. However, most of the fluorescent chemosensors reported still have some limitations, such as long response times, the high limit of detection, and complex synthetic procedures. In this paper, a novel and sensitive fluorescent probe based on Cu2+ modulated polydihydroxyphenylalanine nanoparticles (PDOAs) has been developed for the detection of glyphosate pesticides. The fluorescence of PDOAs can be effectively quenched by Cu2+ through the dynamic quenching process, which was confirmed by the time-resolved fluorescence lifetime analysis. In the presence of glyphosate, the fluorescence of the PDOAs-Cu2+ system can be effectively recovered due to the higher affinity of glyphosate for Cu2+, and thus released the individual PDOAs. Due to the admirable properties such as high selectivity to glyphosate pesticide, "turn on" fluorescence response, and ultralow detection limit of 1.8 nM, the proposed method has been successfully applied for the determination of glyphosate in environmental water samples.

MOF-based surface tailoring the near-infrared luminescence property of gold nanoclusters for ratiometric fluorescence sensing of acetylcholinesterase

Ultrasmall gold nanoclusters (AuNCs) with distinct photoluminescence properties are promising fluorophores for optical applications. However, it remains a great challenge to precisely modulate their near-infrared (NIR) fluorescence properties via a simple yet efficient strategy. Herein, we report a new, surface tailoring strategy for effectively modulating the NIR fluorescence property of AuNCs by harnessing the attractive property of metal-organic frameworks. With zeolite imidazolate frameworks-8 (ZIF-8) as the example, we showed that highly fluorescent AuNCs can be in situ encapsulated into ZIF-8 via a one-pot reaction, yielding AuNCs@ZIF-8 composites with enhanced NIR fluorescence intensity. Particularly, AuNCs@ZIF-8 composites exhibit an emission maximum at 830 nm, which is 190 nm longer than the emission wavelength of AuNCs without the ZIF-8 encapsulation. Mechanism studies based on fluorescence decay spectra and X-ray photoelectron spectroscopy revealed that ZIF-8 mainly poses a significant effect on the ligand-metal charge transfer process of AuNCs due to the confinement effect. By incorporating Rhodamine B (RhB) into NIR-emitting AuNCs@ZIF-8 composites, we established a ratiometric fluorescent sensing platform for the detection of acetylcholinesterase (AChE) based on the turn-on fluorescence response of AuNCs towards thiolates. The AuNCs@ZIF-8/RhB system exhibits a high sensitivity with a detection limit of 0.4 mU/mL and good specificity towards common biological interferences. Moreover, the present ratiometric fluorescence assay shows good recovery for AChE detection in human serum, suggesting its potential application for real sample analysis. Finally, the fluorescence response mechanism of both AuNCs@ZIF-8 and AuNCs towards AChE is elucidated by fluorescence lifetime analysis and density functional theory (DFT) calculation. This study demonstrates the feasibility of modulating the NIR fluorescence property of AuNCs by MOF-based surface tailoring strategy towards enhanced biosensing application.

Selective determination of ellagic acid in aqueous solution using blue-green emissive copper nanoclusters

Development of beneficial sensors to analyze ellagic acid concentrations is of great importance for food safety and human health. Herein, a facile and fast fluorescent probe was carried out for the excellently selective and sensitive measurement of ellagic acid in real samples through histidine protected copper nanoclusters (histidine@Cu NCs) as a nanosensor. This as-developed histidine@Cu NCs were performed through UV–vis absorption spectroscopy, fluorescence spectroscopy, transmission electron microscopy, Fourier-transform infrared spectroscopy, X-ray photoelectron spectroscopy and fluorescence lifetime analysis. The TEM image revealed that this nanomaterial had spherical features with the average diameter of 2.5 ± 0.05 nm. The blue-green fluorescence of this Cu NCs was found under the UV light. Meanwhile, the maximum excitation and emission wavelength were located at 387 nm and 488 nm. After addition of ellagic acid, the fluorescence of histidine@Cu NCs was slowly weakened with excellent linear range of 0.5–300 μM and detection limit of 0.077 μM. The fluorescence weakening mechanism of this nanosensor were attributed to the inner filter effect (IFE) and static quenching. Finally, this as-established analysis platform was successfully employed to measure ellagic acid in real samples.

Multiphoton FLIM Analyses of Native and UVA-Modified Synthetic Melanins.

To better understand the impact of solar light exposure on human skin, the chemical characterization of native melanins and their structural photo-modifications is of central interest. As the methods used today are invasive, we investigated the possibility of using multiphoton fluorescence lifetime (FLIM) imaging, along with phasor and bi-exponential fitting analyses, as a non-invasive alternative method for the chemical analysis of native and UVA-exposed melanins. We demonstrated that multiphoton FLIM allows the discrimination between native DHI, DHICA, Dopa eumelanins, pheomelanin, and mixed eu-/pheo-melanin polymers. We exposed melanin samples to high UVA doses to maximize their structural modifications. The UVA-induced oxidative, photo-degradation, and crosslinking changes were evidenced via an increase in fluorescence lifetimes along with a decrease in their relative contributions. Moreover, we introduced a new phasor parameter of a relative fraction of a UVA-modified species and provided evidence for its sensitivity in assessing the UVA effects. Globally, the fluorescence lifetime properties were modulated in a melanin-dependent and UVA dose-dependent manner, with the strongest modifications being observed for DHICA eumelanin and the weakest for pheomelanin. Multiphoton FLIM phasor and bi-exponential analyses hold promising perspectives for in vivo human skin mixed melanins characterization under UVA or other sunlight exposure conditions.

Fluorescence lifetime analysis of smFRET with contribution of PIFE on donor and acceptor.

Single-molecule fluorescence resonance energy transfer (FRET) is a powerful technique based on dipole-dipole interaction between donor and acceptor fluorophores to observe inter- and intra-molecular dynamics in real time with sensitivity to macro-molecular distances (∼2.5-10 nm). That said, some fluorophores have an inherent characteristic known as protein induced fluorescence enhancement (PIFE). PIFE is a photo-physical feature of dyes undergoing cis-trans transitions and occurs for protein-dye interactions closer than 3 nm where FRET is less sensitive. Here, the challenge is uncoupling the PIFE effect in the FRET data. Ignoring the PIFE effect in the analysis of the FRET data may lead to misinterpretation of the system under investigation. As a solution to this problem, we develop a computational framework based on Bayesian statistics to analyze the fluorescence lifetime signals of the donor and acceptor channels which allows us to uncouple the PIFE effects from the FRET. Our framework can extract any changes in the FRET efficiency simultaneously with any changes in the fluorescence lifetimes of the donor and acceptor due to the PIFE effect. In addition, our framework can provide other parameters, such as the donor and acceptor excitation rates, background photon rates, and detectors’ cross-talk ratios. Our framework extracts all these parameters by analyzing a single photon arrival time trace with only a few thousand photons.