We report on a new method for sensing trace oxygen in the gas phase. It is based on the extreme efficiency of the quenching of the thermally activated delayed fluorescence of isotopically enriched carbon-13 fullerene C(70) ((13)C(70)). This fullerene was dissolved in polymer matrixes of varying oxygen permeability, viz., polystyrene (PS), ethyl cellulose (EC) and an organically modified silica gel ("ormosil"; OS). The sensor films (5-10 μm thick), on photoexcitation at 470 nm, display a strong delayed photoluminescence with peaks between 670 and 700 nm. Its quenching by molecular oxygen was studied at 25 and 60 °C and at concentrations from zero up to 150 ppmv of oxygen in nitrogen. The rapid lifetime determination (RLD) method was applied to determine oxygen-dependent lifetimes and for fluorescence lifetime imaging of oxygen. The lower limits of detection (at 1% quenching) vary with the polymer used (EC ∼250 ppbv, OS ∼320 ppbv, PS ∼530 ppbv at 25 °C) and with temperature. The oxygen sensors reported here are the most sensitive ones described so far.
Current practical methods for finding the equilibrium dissociation constant, K d ,o fp rotein-small molecule complexes have inherent sources of inaccuracy.I ntroduced here is "accurate constant via transient incomplete separation" (ACTIS), which appears to be free of inherent sources of inaccuracy.C onceptually,as hort plug of the pre-equilibrated protein-small molecule mixture is pressure-propagated in ac apillary,c ausing fast transient incomplete separation of the complex from the unbound small molecule.Asuperposition of signals from these two components is measured near the capillary exit and used to calculate af raction of unbound small molecule,which,inturn, is used to calculate K d . Herein the validity of ACTIS is proven theoretically,i ts accuracy is verified by computer simulation, and its practical use is demonstrated. ACTIS has the potential to become ar eference-standardm ethod for determining K d values of protein-small molecule complexes.Reversible binding of proteins (P) to small-molecule ligands (L) plays an important role in the regulation of cellular processes. [1] In addition, most therapeutic targets are proteins, [2] and drugs are developed to form stable PL complexes with them:Complex stability is characterized by the equilibrium dissociation constant K d ,which is defined as:
Robust
and accurate analysis of cell-population heterogeneity is
challenging but required in many areas of biology and medicine. In
particular, it is pivotal to the development of reliable cancer biomarkers.
Here, we prove that cytometry of reaction rate constant (CRRC) can
facilitate such analysis when the kinetic mechanism of a reaction
associated with the heterogeneity is known. In CRRC, the cells are
loaded with a reaction substrate, and its conversion into a product
is followed by time-lapse fluorescence microscopy at the single-cell
level. A reaction rate constant is determined for every cell, and
a kinetic histogram “number of cells versus the rate constant”
is used to determine quantitative parameters of reaction-based cell-population
heterogeneity. Such parameters include, for example, the number and
sizes of subpopulations. In this work, we applied CRRC to a reaction
of substrate extrusion from cells by ATP-binding cassette (ABC) transporters.
This reaction is viewed as a potential basis for predictive biomarkers
of chemoresistance in cancer. CRRC proved to be robust (insensitive
to variations in experimental settings) and accurate for finding quantitative
parameters of cell-population heterogeneity. In contrast, a typical
nonkinetic analysis, performed on the same data sets, proved to be
both nonrobust and inaccurate. Our results suggest that CRRC can potentially
facilitate the development of reliable cancer biomarkers on the basis
of quantitative parameters of cell-population heterogeneity. A plausible
implementation scenario of CRRC-based development, validation, and
clinical use of a predictor of ovarian cancer chemoresistance to its
frontline therapy is presented.
Graphite oxide was characterized by pH dependent excitation-emission matrices from 300 to 500 nm in excitation and from 320 to 600 nm in emission which reveal the presence of two pH steps. These are assigned to the presence of carboxy groups and phenolic hydroxy groups, respectively. Fluorescence is strongest at 470 nm excitation and 555 nm emission. The fluorescence intensity is a function of pH but not of temperature, and is not quenched by oxygen.
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