On characterization of probability distributions by means of independent statistics

Summary The probability density functions f_k(x_k)=A_k|x_k|^p _k−1 e^−aφ _k(x_k) of independent random variables x₀, x₁, ..., x_n, are characterized by independence of two functions of them. Entrata in Redazione il 12 aprile 1969.     

Distributions of fluorescence decay times for synthetic melittin in water-methanol mixtures and complexed with calmodulin,troponin C,and phospholipids

Frequency-domain measurements of the intensity decays of melittin were used to recover the distribution of decay times displayed by its single tryptophan residue. Melittin was examined in the monomeric random coil state (water), in the monomeric -helical state (water-methanol), in the tetrameric state, and with 6M guanidine hydrochloride. In the presence of denaturant, where melittin is expected to be devoid of secondary structure, we observed a narrow distribution of lifetimes, similar to a double-exponential decay. In water the intensity decay of melittin was found to be described better by the distribution of decay times, which became progressively wider as the amount of -helix was increased by the methanol cosolvent or upon formation of the -helical tetrameric state. We also examined the intensity decays of melittin when complexed with calmodulin, troponin C, or lipid vesicles of 1-palmitoyl-2-oleyl-l--phosphatidylcholine (POPC). The lifetime distributions of the complexes with lipid were comparable to those observed in methanol-water, suggesting a similarity of the structure and/or dynamics of the environment surrounding the tryptophan residue. A broad lifetime distribution was observed for the melittin-calmodulin complex, suggesting a rigid structure and/or heterogeneity in the form of the complex. The lifetime distribution of the melittin-troponin C complex was more narrow, suggesting a more uniform structure, at least in the region surrounding the tryptophan residue. These results demonstrate that the lifetime distributions of a single tryptophan protein can be a sensitive indicator of the conformational heterogeneity and dynamics of proteins.     

Increased intensities of YOYO-1-labeled DNA oligomers near silver particles

DNA detection is usually performed using fluorescence probes. Using a DNA oligomer stained with the widely used dye 1,1'-[1,3-propanediylbis[(dimethylimino)-3,1-propanediyl]]bis[4-[(3-methyl-2(3H)-benzoxazolylidene)methyl]]-quinolinum tetraiodide (YOYO-1), we show that a substrate containing silver particles can lead to a greater than 10-fold increase in the fluorescence intensity. Proximity to silver particles also increases the photostability of YOYO-1-DNA. These results suggest that substrates or gels containing silver particles may be used for increased sensitivity in DNA detection.     

PICOSECOND RESOLUTION OF INDOLE ANISOTROPY DECAYS AND SPECTRAL RELAXATION BY 2 GHz FREQUENCY-DOMAIN FLUOROMETRY

Abstract— We used frequency-domain fluorescence spectroscopy to measure rotational diffusion and time-resolved emission spectra of indole in methanol on the picosecond timescale. The indole emission was quenched by acrylamide to allow measurements to the instrumental limit of 2 GHz and to eliminate emission from the longer-lived indole molecules, which can no longer provide information on the picosecond (ps) processes. The resolution was adequate to measure rotational correlation times as short as 8 ps at 80†C, and spectral relaxation times as short as 16 ps at 5†C.     

Evaluation of aging characteristics of bituminous roof coverings by thermo-mechanical analysis (TMA)

Four bituminous roof coverings have been subjected to various aging procedures including heat, UV irradiation and temperature/rain cycling. A specially modified TMA technique has been used for evaluation, depending on measurements of softening temperature. A comparison with mechanical test data has also been made. Some conclusions have been drawn with respect to influence of heat and UV radiation on the rate of degradation.     

Multiphoton Ligand-Enhanced Excitation of Lanthanides

We describe multiphoton excitation of the lanthanides europium (Eu³⁺) and terbium (Tb³⁺) when these ions are complexed with nucleic acids, proteins, and fluorescent chelators. In all cases excitation occurs by multiphoton absorption of the sensitizers. For the nucleotide GDP and an oligonucleotide with several guanines, the sensitized emission of Tb³⁺ excited at 776 nm indicated a three-photon process. For Tb³⁺ bound to the wild-type troponin C and a single tryptophan mutant (26W), excitation at 794 nm was also close to a three-photon process. For lanthanide chelators containing various sensitizers, we observed three-photon excitation in the case of methyl anthranilate, a mixuture of two- and three-photon excitation for carbostyril 124, and a two-photon process with a coumarin derivative. In the case of coumarin-sensitized emission of Eu³⁺ varied from a two- to a three-photon process at wavelengths ranging from 780 to 880 nm. The sensitized luminescence also shows significantly higher photostability compared to the fluorescence from the organic fluorophores alone. These results suggest the use of multiphoton-induced sensitized lanthanide fluorescence in biochemistry and cellular imaging.On leave from the Institute of Experimental Physics     

Elimination of autofluorescence in fluorescence correlation spectroscopy using the AzaDiOxaTriAngulenium (ADOTA) fluorophore in combination with time-correlated single-photon counting (TCSPC)

Fluorescence correlation spectroscopy (FCS) is a frequently applied technique that allows for the precise and sensitive analysis of molecular diffusion and interactions. However, the potential of FCS for in vitro or ex vivo studies has not been fully realized due in part to artifacts originating from autofluorescence (fluorescence of inherent components and fixative-induced fluorescence). Here, we propose the azadioxatriangulenium (ADOTA) dye as a solution to this problem. The lifetime of the ADOTA probe, about 19.4 ns, is much longer than most components of autofluorescence. Thus, it can be easily separated by time-correlated single-photon counting methods. Here, we demonstrate the suppression of autofluorescence in FCS using ADOTA-labeled hyaluronan macromolecules (HAs) with Rhodamine 123 added to simulate diffusing fluorescent background components. The emission spectrum and decay rate of Rhodamine 123 overlap with the usual sources of autofluorescence, and its diffusion behavior is well known. We show that the contributions from Rhodamine 123 can be eliminated by time gating or by fluorescence lifetime correlation spectroscopy (FLCS). While the pairing of ADOTA and time gating is an effective strategy for the removal of autofluorescence from fluorescence imaging, the loss of photons leads to erroneous concentration values with FCS. On the other hand, FLCS eliminates autofluorescence without such errors. We then show that both time gating and FLCS may be used successfully with ADOTA-labeled HA to detect the presence of hyaluronidase, the overexpression of which has been observed in many types of cancer.