Fluorescence Lifetime Techniques Research Articles

Unlocking the luminescent potential of Pr3+/Yb3+ Co-doped Y2Mo4O15 for advanced thermometry applications

In this study, Pr3+/Yb3+ co-doped Y2Mo4O15 phosphor was successfully synthesized using the traditional citric acid-assisted sol-gel method, with the aim of exploring its potential for optical thermometry applications. The structural and morphological characteristics of the synthesized sample were comprehensively analyzed through X-ray powder diffraction (XRD) and scanning electron microscopy (SEM). Photoluminescence and temperature-dependent luminescence properties were systematically investigated within a temperature range of 298 K to 508 K, employing 473 nm excitation. To assess the temperature sensing capabilities of the phosphors, we utilized both the fluorescence intensity ratio (FIR) and fluorescence lifetime (FL) techniques. Optical temperature measurements were conducted through both thermal and non-thermal coupling methods. Our findings revealed that the FIR (I547/I490) exhibited an outstanding maximum sensitivity of 1.27% K−1, accompanied by remarkably low-temperature uncertainty (δT) values ranging from 0.04 to 0.83 K. This outstanding performance makes it the optimal choice for temperature sensing. Additionally, when employing FL thermometry, the maximum sensitivity (Sr) was determined to be 0.21% K−1 at 538 K. These results strongly indicate the significant potential of Y2Mo4O15: Pr3+/Yb3+ in the realm of optical thermometry, offering promising opportunities for precise and reliable temperature measurements.

Rapid and ratiometric fluorescent detection of phosgene by a red-emissive ESIPT-based-benzoquinolone probe

Phosgene is a highly toxic gas that poses a serious threat to human health and public safety. Therefore, it is of great importance to develop an available detection method enabling on-the-spot measurement of phosgene. In this paper, we report a novel ESIPT fluorescent probe for phosgene detection based on quinolone fluorophore. This probe exhibits rapid response (in 10 s), stable signal output (last for 10 min), high sensitivity (LOD ∼ 6.7 nM), and distinct emission color change (red to green) towards phosgene. The sensing mechanism was investigated by using 1H NMR, HRMS and fluorescence lifetime techniques, confirming that the amidation reaction between phosgene and quinolone effectively suppressed the ESIPT process of probe. Eventually, this probe was fabricated into polymer nanofibers by electrospinning and successfully employed to monitor gaseous phosgene with high specificity. This work provided a promising analytical tool for rapid and ratiometric detection of phosgene both in solution and in the gas phase.

Comparing different modalities of time-resolved two-photon fluorescence measurements for FRET analysis

Different modalities of time-resolved fluorescence lifetime techniques are typically used for Förster resonance energy transfer (FRET) studies in vivo and in solution and have been used to measure macromolecular processes and interactions. Here, we examine the sensitivity of the estimated FRET efficiency and donor-acceptor distance to different modalities of time-resolved two-photon (2P) fluorescence measurements, experimental design, and data analysis. As a model, we used the biosensor (mTurquoise-linker-mCitrine), where mTurquoise-mCitrine serves as the FRET pair tethered with a flexible linker.

Fluorescence quantum yields and lifetimes of annatto aqueous solutions dependent on hydrogen potential: Applications in adulterated milk

This work reports the thermo-optical and photophysical properties of annatto extracted from the seeds of the tropical shrub Bixa Orellana L. Aqueous natural dyes were extracted using different amounts of NH4OH solution (4 M) and separated into two groups of annatto samples extracted using ambient temperature (G1Amb) and heated (98 °C, G2Heat) alkaline solutions. Thermal lens (TL) and time-resolved fluorescence lifetime (TRFL) techniques were applied to determine the fluorescence quantum yield (η) and lifetime (τ) for G1Amb and G2Heat colorant solutions with different hydrogen potentials (pHs) ranging from 5.4 to 11.5. Fluorescence spectra corroborate the η results for natural dye extracted at different pH values. Additionally, the annatto dyes were tested as possible colorimetric and/or fluorimetric pH-biosensor probes in adulterated milk at different pHs.

Compact electron spin resonance skin oximeter: Properties and initial clinical results

Skin oxygen level is of significance for the diagnosis and treatment of many clinical problems, such as chronic wounds and diabetic foot ulcers. Furthermore, skin oxygen levels can be correlated to arterial oxygen partial pressure, thereby revealing potentially dangerous conditions such as hyperoxia (too much oxygen), which may occur in ventilated neonates. Traditionally, skin oxygen levels are measured using electrochemical methods and, more recently, also by fluorescence lifetime techniques. These approaches suffer from several drawbacks, rendering them suboptimal. The purpose of this work is to develop an electron spin resonance (ESR) -based method for monitoring oxygen partial pressure (pO2 ) in skin tissue. A compact sensor for pulsed ESR is designed and constructed. Our ESR-based method makes use of a unique exogenous paramagnetic spin probe that is placed on the skin in a special partially sealed sticker, and subsequently measuring its signal with the compact pulsed ESR sensor that includes a miniature magnet and a small S-band (~2.3 GHz) microwave resonator. The inverse of the spin-spin relaxation time (1/T2 ) measured by ESR is shown to be linearly correlated with pO2 levels. The sensor and its matching sticker were tested both in vitro and in vivo (with human subjects). Measured skin pO2 levels reached equilibrium after ~2-3 h and were found to be comparable to those measured by continuous-wave (CW) ESR using a large electromagnet. A compact pulsed ESR sensor with a matching paramagnetic sticker can be used for pO2 monitoring of the skin tissue, similar to large bulky CW ESR systems.

Spectroscopy and molecular docking approach for investigation on the binding of nocodazole to human serum albumin.

The interaction between nocodazole (Nz) and human serum albumin (HSA) under controlled physiological condition (pH7.4) is examined using absorption, emission, fluorescence lifetime (FLT) and circular dichroism (CD) spectroscopic techniques. The binding constant (order of 105M-1) from UV-vis and fluorescence spectroscopy reveals a strong interaction between Nz and HSA. Fluorescence quenching study shows that Nz binds with HSA through static quenching process. It is induced by formation of Nz-HSA complex because the Stern-Volmer quenching constant is inversely correlated with the temperature which is further verified by time-resolved fluorescence spectroscopy. The thermodynamic parameters at different temperatures indicate that the binding process is spontaneous where hydrogen bonding interactions and Van der Waals forces play major roles during the interaction between Nz and HSA. By means of spectroscopy and molecular modeling, we have discovered and interpreted the alteration of the secondary structure of HSA by Nz complexation. Synchronous, three-dimensional fluorescence and CD spectroscopic results reveal that the addition of Nz to HSA affects changes in the micro-environment and conformation of HSA. According to Förster Resonance Energy Transfer (FRET), the binding distance (r) between Nz and residue of HSA is <8nm with excellent energy efficiency. The docking study suggests that nocodazole binds at Domain IIA in the hydrophobic pocket of human serum albumin.

Is Gum Arabic a Good Emulsifier Due to CH...π Interactions? How Urea Effectively Destabilizes the Hydrophobic CH...π Interactions in the Proteins of Gum Arabic than Amides and GuHCl?

The photophysical studies of gum arabic (GA) in the presence of urea, 1,3-dimethylurea (DMU), tetramethylurea (TMU), guanidine hydrochloride (GuHCl), formamide (FA), acetamide (AA), and dimethyl formamide (DMF) were carried out by monitoring the emission, three-dimensional emission contour, and time-correlated fluorescence lifetime techniques. On addition of only 1 × 10–3 M urea, 75.0% of the fluorescence of GA is quenched, while the same occurs in GuHCl at 3.0 M. FA quenched 50% of the fluorescence of GA at 5.0 M. However, DMU, TMU, AA, and DMF resulted in a fluorescence enhancement. The unusual fluorescence trends reveal the existence of CH...π interactions in the proteins of GA. The experimental results and the structural aspects of proteins in GA led us to propose that the aggregation of polyproline helices in GA, through several CH...π interactions, would have a major role to play in the emulsification mechanism of GA.

Fluorescence intensity ratio and lifetime thermometry of praseodymium phosphates for temperature sensing

We report on the temperature sensing properties of varied praseodymium phosphate hosts MI(Na,Li,K)Pr(PO3)4 and PrP5O14. Details about fabrication, materials, and spectroscopic characterization are presented. The fluorescence properties of the praseodymium phosphates have been systematically investigated from room temperature to 363 K under selective laser excitation in the 3P0 multiplet. The temperature sensing measurements were carried out by using fluorescence intensity ratio and fluorescence lifetime techniques. From this systematic study, the thermal resolution potentially achieved has been determined and compared to previous works.

Evaluating Cell Metabolism Through Autofluorescence Imaging of NAD(P)H and FAD.

Optical imaging using the endogenous fluorescence of metabolic cofactors has enabled nondestructive examination of dynamic changes in cell and tissue function both in vitro and in vivo. Quantifying NAD(P)H and FAD fluorescence through an optical redox ratio and fluorescence lifetime imaging (FLIM) provides sensitivity to the relative balance between oxidative phosphorylation and glucose catabolism. Since its introduction decades ago, the use of NAD(P)H imaging has expanded to include applications involving almost every major tissue type and a variety of pathologies. Recent Advances: This review focuses on the use of two-photon excited fluorescence and NAD(P)H fluorescence lifetime techniques in cancer, neuroscience, tissue engineering, and other biomedical applications over the last 5 years. In a variety of cancer models, NAD(P)H fluorescence intensity and lifetime measurements demonstrate a sensitivity to the Warburg effect, suggesting potential for early detection or high-throughput drug screening. The sensitivity to the biosynthetic demands of stem cell differentiation and tissue repair processes indicates the range of applications for this imaging technology may be broad. As the number of applications for these fluorescence imaging techniques expand, identifying and characterizing additional intrinsic fluorophores and chromophores present in vivo will be vital to accurately measure and interpret metabolic outcomes. Understanding the full capabilities and limitations of FLIM will also be key to future advances. Future work is needed to evaluate whether a combination of different biochemical and structural outcomes using these imaging techniques can provide complementary information regarding the utilization of specific metabolic pathways.

Thermal diffusivity and lifetime studies of Styryl 7 dye on DNA-CTMA complex

This study reports the sensing of micro viscosity variation of Styryl 7(LDS 750) dye in different solvents and biological samples through thermal lens and fluorescence lifetime techniques. Fluorescence properties of Styryl 7 dye in different solvents and with DNA-CTMA (deoxyribonucleic acid attached with cetyltrimethyl-ammonium cationic surfactant molecules) were measured using fluorescence lifetime technique. Fluorescence lifetime of Styryl 7 in different solvents are found to increase with the viscosity of the samples. With increase in weight percentage of DNA-CTMA, the fluorescence lifetime increases from 0.17ns to1.47ns. Lifetime studies confirm that Styryl 7 dye is a good fluorescent probe for detecting the micro viscosity of the samples. Thermal lens technique, which is an efficient probe to measure non-radiative relaxation route were used to measure the thermal diffusivity of Styryl 7 in different solvents and with varying weight percentage of DNA-CTMA. Thermal diffusivity of the samples are found to decrease with increase in solvent viscosity and increased weight percentage of DNA-CTMA

Portable fluorescence lifetime spectroscopy system for in-situ interrogation of biological tissues.

Fluorescence spectroscopy and lifetime techniques are potential methods for optical diagnosis and characterization of biological tissues with an in-situ, fast, and noninvasive interrogation. Several diseases may be diagnosed due to differences in the fluorescence spectra of targeted fluorophores, when, these spectra are similar, considering steady-state fluorescence, others may be detected by monitoring their fluorescence lifetime. Despite this complementarity, most of the current fluorescence lifetime systems are not robust and portable, and not being feasible for clinical applications. We describe the assembly of a fluorescence lifetime spectroscopy system in a suitcase, its characterization, and validation with clinical measurements of skin lesions. The assembled system is all encased and robust, maintaining its mechanical, electrical, and optical stability during transportation, and is feasible for clinical measurements. The instrument response function measured was about 300ps, and the system is properly calibrated. At the clinical study, the system showed to be reliable, and the achieved spectroscopy results support its potential use as an auxiliary tool for skin diagnostics.

Note: Rapid measurement of fluorescence lifetimes using SiPM detection and waveform sampling.

In fluorescence spectroscopy and imaging, fluorescence lifetime measurement-assessing the average time fluorophores spend in their excited state before returning to their ground state-offers a number of advantages over quantifying fluorescence intensities that include resistance to photo-bleaching and independence from fluorophore concentration, excitation intensity, and measurement methodology. Despite growing interest, fluorescence lifetime techniques frequently mandate relatively complex instrumentation, slow data acquisition rates, and significant data analyses. In this work, we demonstrate the feasibility of measuring fluorescence lifetimes using off-the-shelf analog silicon photomultipliers and switched-capacitor array waveform sampling techniques, with precision matching that of much larger and more elaborate commercial instruments.

Synthesis and optical properties of Y2Mo4O15 doped by Pr3+

A set of Y2Mo4O15 compounds doped with 0.1–30% Pr3+ was synthesized by conventional high temperature solid state synthesis. The structural, morphological and optical characteristics of the compounds were investigated by powder X-ray diffraction (XRD), scanning electron microscopy (SEM) analysis, thermal quenching (TQ), photoluminescence (PL) and fluorescence lifetime techniques. It turned out that Pr3+ emission spectra are very sensitive to the concentration and temperature in the Y2Mo4O15 host matrix. Samples showed mainly red and deep red luminescence when excited with blue radiation. A severe concentration quenching was observed in samples with higher Pr3+ content due to cross relaxation. Y2Mo4O15:Pr3+ luminescent materials possess rather unique emission spectra, which can be tuned by adopting the appropriate Pr3+ concentration and/or temperature. Thus these materials could be used as luminescent security markers where such properties are desirable.

NADH Auto Fluorescence Reveals New Metabolic Signatures in Yeast and Mammalian Cells

Yeast cells and their extracts were reported to show glycolytic fluctuations with a frequency of several minutes with addition of supplements. There is a distinct phase relation of NADH and other oscillating metabolites and intermediates which can be studied observing the auto fluorescence. New fast and efficient fluorescence lifetime techniques emerged in last two decades accelerated the progress of revealing the true nature of these fluctuations. We address these dynamic changes happening in live yeast systems by imaging them in-vivo time lapse lifetime imaging and correlate it with carbon metabolism reported in mammalian and cancer cells. The similarity between cancer cell metabolic flux and rapidly proliferating yeast cells have recently been of scientific interest to elucidate cancer cell pathways. Our findings shed new light into these metabolic responses with help of auto fluorescence lifetime measurements and live flux measurements. In addition, we aid these observations with the rigorous validation of auto fluorescence lifetime based metabolic studies.

Effects of N-acetyl-L-cysteine-capped CdTe quantum dots on bovine serum albumin and bovine hemoglobin: isothermal titration calorimetry and spectroscopic investigations.

The interactions of N-acetyl-L-cysteine-capped CdTe quantum dots (QDs) with bovine serum albumin (BSA) and bovine hemoglobin (BHb) were investigated by isothermal titration calorimetry (ITC), fluorescence, synchronous fluorescence, fluorescence lifetime, ultraviolet-visible absorption, and circular dichroism techniques. Fluorescence data of BSA-QDs and BHb-QDs revealed that the quenching was static in every system. While CdTe QDs changed the microenvironment of tryptophan in BHb, the microenvironment of BSA kept unchanged. Adding CdTe QDs affected the skeleton and secondary structure of the protein (BSA and BHb). The ITC results indicated that the interaction between the protein (BSA and BHb) and QDs-612 was spontaneous and the predominant force was hydrophobic interaction. In addition, the binding constants were determined to be 1.19 × 10(5) L mol(-1) (BSA-QDs) and 2.19 × 10(5) L mol(-1) (BHb-QDs) at 298 K. From these results, we conclude that CdTe QDs have a larger impact on the structure of BHb than BSA.

Multimodal in vivo imaging of oral cancer using fluorescence lifetime, photoacoustic and ultrasound techniques

This work reports a multimodal system for label-free tissue diagnosis combining fluorescence lifetime imaging (FLIm), ultrasound backscatter microscopy (UBM), and photoacoustic imaging (PAI). This system provides complementary biochemical, structural and functional features allowing for enhanced in vivo detection of oral carcinoma. Results from a hamster oral carcinoma model (normal, precancer and carcinoma) are presented demonstrating the ability of FLIm to delineate biochemical composition at the tissue surface, UBM and related radiofrequency parameters to identify disruptions in the tissue microarchitecture and PAI to map optical absorption associated with specific tissue morphology and physiology.

Fluorescence lifetime techniques in medical applications.

This article presents an overview of time-resolved (lifetime) fluorescence techniques used in biomedical diagnostics. In particular, we review the development of time-resolved fluorescence spectroscopy (TRFS) and fluorescence lifetime imaging (FLIM) instrumentation and associated methodologies which allow in vivo characterization and diagnosis of biological tissues. Emphasis is placed on the translational research potential of these techniques and on evaluating whether intrinsic fluorescence signals provide useful contrast for the diagnosis of human diseases including cancer (gastrointestinal tract, lung, head and neck, and brain), skin and eye diseases, and atherosclerotic cardiovascular disease.

Saddle Distortion of a Sterically Unhindered Porphyrin Ring in a Copper Porphyrin with Electron-Donating Substituents

Two crystalline porphyrins, CuT(4'-OMePh)P (1) and H2T(4'-OMePh)P (2) (T(4'-OMePh)P(2-) = 5, 10, 15,20-tetrakis(4-methoxyphenyl)-21H,23H-porphyrin dianion), have been synthesized and characterized by a single-crystal X-ray diffraction. Compound 1 is crystallized in the orthorhombic system with a space group of Pna21, but compound 2 is crystallized in the monoclinic system with a space group of P2/c. Compound 1 is characterized as an isolated structure with a saddle-distorted nonplanar porphyrin macrocycle and an embedded cupric ion coordinating to four pyrrole nitrogen atoms. Nonmetalated compound 2 also displays an isolated structure, but the macrocycle of porphyrin is close to a perfect plane. The molecules in 2 are interconnected through five C-H · · · π hydrogen bonding interactions to yield a 3-D supramolecular network. However, the molecules in 1 are interlinked via five C-H · · · π interactions and two C-H · · · O hydrogen bonding interactions to yield a more complex 3-D supramolecular motif. The two more C-H · · · O hydrogen bonding interactions are attributed to the distortion of porphyrin macrocycle, resulting from the metalation. The metalation brought changes not only in the crystal structures and supramolecular motifs but also in the properties. The photophysical and redox properties of 1 and 2 in solution have also been studied by steady-state absorption and fluorescence, cyclic voltammetry, fluorescence lifetime and nanosecond transient absorption techniques.

Fluorescence lifetime in cardiovascular diagnostics.

We review fluorescence lifetime techniques including time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) and fluorescence lifetime imaging microscopy (FLIM) instrumentation and associated methodologies that allow for characterization and diagnosis of atherosclerotic plaques. Emphasis is placed on the translational research potential of TR-LIFS and FLIM and on determining whether intrinsic fluorescence signals can be used to provide useful contrast for the diagnosis of high-risk atherosclerotic plaque. Our results demonstrate that these techniques allow for the discrimination of important biochemical features involved in atherosclerotic plaque instability and rupture and show their potential for future intravascular applications.

Fluorescence Study of Inclusion Complexes between Star-Shaped Cholic Acid Derivatives and Polycyclic Aromatic Fluorescent Probes and the Size Effects of Host and Guest Molecules

Star-shaped host molecules containing two, three, and four cholic acid moieties have been used to form inclusion complexes with polycyclic aromatic hydrocarbon probes (guests) varying in size from four (pyrene) to five (benzo(e)pyrene) and seven aromatic rings (coronene) and investigated by steady-state fluorescence measurements and fluorescence lifetime techniques. The results indicated that these hydrophobic guest probes prefer to locate in the hydrophobic cavities formed by the host molecules in an aqueous solution. Further studies showed that the stoichiometric ratios of the complexes depended on the relative size of both the host and the guest. The complexes of 1:1 ratio (guest:host) were formed between pyrene and the host molecules of different sizes, while the complexes of 1:2 ratio (guest:host) were found for coronene in all cases. For benzo(e)pyrene with an intermediate size, the complexes with 1:1 and 1:2 ratios (guest:host) were formed depending on the relative sizes of the host molecules. The stability of the inclusion complexes was observed to change with the solvent polarity, indicative of an adaptation of the hydrophobicity of the host pockets to the polarity of the solvent. The formation of the complexes was driven by the solvophobic interactions.