Label-free sensing of cells with fluorescence lifetime imaging: the quest for metabolic heterogeneity

Molecular, morphological and physiological heterogeneity is the inherent property of cells, which governs differences in their response to external influence. The tumor cells metabolic heterogeneity is of a special interest due to its clinical relevance to the tumor progression and therapeutic outcomes. Rapid, sensitive and non-invasive assessment of metabolic heterogeneity of cells is of a great demand for biomedical sciences. Fluorescence lifetime imaging (FLIM), which is an all-optical technique is an emerging tool for sensing and quantifying cellular metabolism by measuring fluorescence decay parameters (FDPs) of endogenous fluorophores, such as NAD(P)H. To achieve the accurate discrimination between metabolically diverse cellular subpopulations, appropriate approaches to FLIM data collection and analysis are needed. In this report, the unique capability of FLIM to attain the overarching goal of discriminating metabolic heterogeneity has been demonstrated. This has been achieved using a novel approach to data analysis based on the non-parametric analysis, which revealed a much better sensitivity to the presence of metabolically distinct subpopulations as compare more traditional approaches of FLIM measurements and analysis. The new approach was further validated for imaging cultured cancer cells treated with chemotherapy. Those results pave the way for an accurate detection and quantification of cellular metabolic heterogeneity using FLIM, which will be valuable for assessing therapeutic vulnerabilities and predicting clinical outcomes.

Efficient near-infrared quantum cutting by cooperative energy transfer in Bi3TeBO9:Nd3+ phosphors

An efficient near-infrared quantum cutting process by cooperative down-conversion of active Bi3+ and Nd3+ ions was demonstrated in Bi3TeBO9:Nd3+ phosphors. In particular, the near-infrared emission of Nd3+ ions enhanced by Bi3+ ions of a series of novel Bi3TeBO9:Nd3+ microcrystalline powders doped with Nd3+ ions in various concentrations was investigated. In order to investigate the luminescent properties of BTBO:Nd3+ powders, the excitation and emission spectra and the fluorescence decay time were measured and analyzed. In particular, the emission of Bi3TeBO9:Nd3+ at 890 and 1064 nm was excited at 327 nm (via energy transfer from Bi3+ ions) and at 586.4 nm (directly by Nd3+ ions). The highest intensity emission bands in near-infrared were detected in the spectra of Bi3TeBO9:Nd3+ doped with 5.0 and 0.5 at.% of Nd3+ ions upon excitation in ultraviolet and visible spectral range, respectively. The fluorescence decay lifetime monitored at 1064 nm for Bi3TeBO9:Nd3+ powders shows the single- or double-exponential character depending on the concentrations of Nd3+ ions. The possible mechanisms of energy relaxation after excitation Bi3TeBO9:Nd3+ powders in ultraviolet or visible spectral range were discussed. The investigated Bi3TeBO9:Nd3+ phosphors efficiently concentrate the ultraviolet/visible radiation in the near-infrared spectral range and can be potentially used as effective spectral converters. Graphical abstract

Mn4+ non-equivalent doped fluoride phosphor towards a short fluorescence decay time for backlighting

The non-equivalent doping of Mn4+ in the red-emitting fluoride phosphors effectively shortens the fluorescence lifetime. Herein, we successfully synthesized Rb2NaInF6:Mn4+ phosphors by an ion-exchange method. The compensation mechanism of Mn4+...

Analyses of energy transfer of Cr3+ and Nd3+ co-doped Y3Al5O12 ceramic powders at the 4T1 level of Cr3+ ion excitation

The concentration dependence of energy transfer from Cr3+ to Nd3+ at the 4T1 level excitation in Nd/Cr:YAG was investigated by the fluorescence decay curves of Cr3+ and Nd3+ for Nd/Cr:YAG and Cr:YAG ceramic powders in the Cr3+ concentration range of 0.1 to 6.0 mol%. The energy transfer process between Cr3+ and Nd3+ at the 4T1 level excitation is tried to explain using a rate equation that assumes energy transfer from the 2E–4T2 level to Nd3+ on the basis of dipole–dipole interactions, the same as the 4T2 level excitation. In conclusion, the energy excited to the 4T1 level will relax non-radiatively to the 2E–4T2 level and then transfer to Nd3+. It is presumed there will be no direct transfer from the 4T1 level to Nd3+. Our rate equations will be useful when simultaneously exciting the 4T1 and 4T2 levels of Cr3+ in Nd/Cr:YAG using broadband pumping sources.

Dosimeter Based on YAG: Ce Phosphor via Sol-Gel Method for Online X-ray Radiation Monitoring

This paper focuses on the preparation of cerium-doped yttrium aluminum garnet (YAG: Ce) powder with several concentration gradients via the sol-gel method by detecting its structural characteristics via X-ray diffraction (XRD) patterns and scanning electron microscope (SEM) to verify the generation of a complete crystal phase and evenly distributed nanopowder. On this basis, the luminescence characteristics of Ce3+ are explored, the mechanism and model are discussed based on the spectra, and the ideal doping concentration was obtained by comparing the luminescence intensity along with the fluorescence quenching theory and fluorescence decay spectra of samples with different doping concentrations. Several radiation dosimeters based on YAG: Ce phosphors were made; the online radiation monitoring function was realized under the exposure of a standard X-ray source; the repeatability, accuracy, and sensitivity of the system were verified by experiments; and the factors affecting dosimeter response are discussed. This paper verifies the possibility of adhibiting YAG: Ce fluorescent powder for online X-ray monitoring, and lays the foundation for further research.

Analysis of Fluorescence Decay Kinetics of Indocyanine Green Monomers and Aggregates in Brain Tumor Model In Vivo

Spectroscopic approach with fluorescence time resolution allows one to determine the state of a brain tumor and its microenvironment via changes in the fluorescent dye’s fluorescence lifetime. Indocyanine green (ICG) is an acknowledged infra-red fluorescent dye that self-assembles into stable aggregate forms (ICG NPs). ICG NPs aggregates have a tendency to accumulate in the tumor with a maximum accumulation at 24 h after systemic administration, enabling extended intraoperative diagnostic. Fluorescence lifetime analysis of ICG and ICG NPs demonstrates different values for ICG monomers and H-aggregates, indicating promising suitability for fluorescent diagnostics of brain tumors due to their affinity to tumor cells and stability in biological tissue.

A study of the subdiffusion of small molecules in charged polyelectrolyte multilayers

A theoretical approach has been developed here to describe the slow diffusion of small charged molecules of sodium dithionite (S2O42−) in polyelectrolyte multilayers (PEMs) composed of polyallylamine hydrochloride (PAH) and polystyrene sulfonate (PSS), which is demonstrated here to be a case of subdifussion. Diffusion is measured experimentally by recording the quenching of the fluorescence of (7-nitrobenz-2-oxa-1,3-diazol-4yl) amino (NBD) labelled PAH layers assembled on silica particles by flow cytometry. NBD is reduced when it encounters dithionite leading to the disappearance of the fluorescence. The fluorescence decay curves show a slow diffusion of dithionite, that does not follow classical Fickean law. Dithionite diffusion in the PEMs is shown to be a non-Markovian process and the slow diffusion can be described via diffusion equations with fractional time derivatives. Results are explained assuming subdifussion of dithionite in the PEMs, as a result of the trapping of the negatively charged dithionite in the positively charged layers of PAH.

Effects of size and dimension in optic relaxation in Rb vapor

We report on an experiment in which the fluorescence decay time of 5P levels of Rb atoms in a coated vapor cell exceeds several of milliseconds that many orders of magnitude longer than normal decay time of excited of rubidium atoms. This effect has been observed by using the pump laser with fixing frequency and the periodically scan the probing laser frequency of which overlap all Rb hyperfine linens of 5P-5S transition. Long time fluorescence decay take place in wide part of the spherical high quality coated cell. In cylindrical part of the cell or in uncoated cell the long time fluorescence decay was disappeared.

Altering the distribution of excited-state lifetimes in aminated GFP chromophores by Ag nanohole arrays

Fluorescence of the modified GFP chromophore diethyl-ABDI-BF2 dispersed in PMMA matrix is studied on top of glass, continuous and perforated optically thin silver films. In polymer, the fluorescence decay kinetics becomes non-exponential and can be described by the distribution of rate constants. The results demonstrate shortening of the excited state lifetime in the presence of silver and broadening of the lifetime distribution caused by the nanoholes.