Comparison of FÖrster-Resonance-Energy-Transfer Acceptors for Tryptophan and Tyrosine Residues in Native Proteins as Donors

Homogenous bioaffinity analysis with tryptophan/tyrosine residues in native proteins as FÖrster-resonance-energy-transfer (FRET) donors is feasible when suitable fluorophors can act as FRET acceptors in ligands (FRET probes) and FRET efficiency in complexes of proteins and FRET probes is high enough. In complexes of proteins and FRET probes, suitable acceptors should have excitation peaks around 335 nm and high rotation freedom, are preferred to have sufficient quantum yields and excitation valleys around 280 nm. In protein binding sites mimicked with mixtures of neutral phosphate buffer and organic solvents, quantum yields of candidate acceptors are altered inconsistently but their excitation peaks show tiny changes. Fluorophores as acceptors in such FRET probes are buried inside glutathione-S-transferase and have low rotation freedom, but are localized on streptavidin surface and display high rotation freedom; FRET efficiency in complexes of streptavidin and its FRET probes is much stronger than that in complexes of glutathione-S-transferase and its FRET probes. Specially, the quantum yield is about 0.70 for free 1-naphthylamine probe in neutral phosphate buffer, about 0.50 for 1-naphthylamine probe bound by streptavidin, and about 0.15 for that bound by glutathione-S-transferase. The quantum yield is about 0.06 for free dansylamide probe, about 0.11 for dansylamide probe bound by streptavidin and about 0.27 for that bound by glutathione-S-transferase. Therefore, 1-naphthylamine and dansylamide are effective acceptors when they localize on surfaces of complexes of proteins and FRET probes.     

Electronic properties of Hf2X intermetallic compounds (X = Al, Si, Ni, Ga and Ge)

Using ¹⁰⁰Pd/¹⁰⁰Rh probes, perturbed angular correlation measurements were performed to study Pd-related defects in Si as a function of dopant concentration and dopant type. Pd-vacancy and Pd-B complexes were identified by their characteristic electric field gradients in highly doped n- and p-type Si, respectively. Both pairs exhibited a T^3/2 temperature dependence of their electric field gradients.     

Long-lifetime metal-ligand complexes as luminescent probes for DNA

We examined the intensity and anisotropy decays of DNA labeled with two ruthenium metalligand complexes, [Ru(bpy)₂(dppz)]²⁺ and [Ru(phe)₂(dppz)]²⁺. Both complexes display high emission anisotropies in the absence of rotational diffusion, making them suitable probes for rotational motions. When bound to DNA, these complexes display decay times as long as 294 ns, providing long-lived probes of DNA dynamics. The decay times of both complexes were rather insensitive to dissolved oxygen. We examined anisotropy decays of these complexes bound to B-form DNA. The anisotropy decays revealed correlation times near 10, 50, and several hundred nanoseconds, suggesting that these probes are sensitive to a wide range of DNA motions. The use of metalligand complexes should allow resolution of both the torsional and bending motions of DNA, the latter of which has been mostly inaccessible using shorter-lived fluorescent probes bound to DNA. Dedicated to Professor Robert F. Steiner upon his retirement     

Membrane Dipole Potential as Measured by Ratiometric 3-Hydroxyflavone Fluorescence Probes: Accounting for Hydration Effects

We previously applied the electrochromic modulation of excited-state intramolecular proton-transfer (ESIPT) reaction for the design of novel 3-hydroxyflavone (3-HF) derivatives as fluorescent probes for measuring the dipole potential, Ψ_D, in lipid bilayers (Klymchenko et al., Proc. Natl. Acad. Sci. USA, 2003, 100, 11219). In the present work, this method was revisited to take into account the influence of the bilayer hydration on the emission ratiometric response of 3-HF probes. For this reason, it was necessary to deconvolute the whole fluorescence spectra into three bands corresponding to the non H-bonded forms, normal N^* and tautomer T^* forms, both participating to the ESIPT reaction, and to the H-bonded H–N^* form, excluded from this reaction. This allowed us to determine the pure N^*/T^* intensity ratio, without any contribution from the H–N^* form emission depending essentially on the bilayer hydration. This new approach allowed us to confirm the correlation we obtained between the response of 3-HF probes on dipole potential modifications and the corresponding response of the reference fluorescent probe di-8-ANEPPS, thus further confirming the potency of 3-HF probes as excellent emission ratiometric probes to measure dipole potential in lipid membranes.     

Polarized emission from a rhenium metal-ligand complex

We report the first observation of polarized emission from a rhenium-phenanthroline complex, Re(CO)₃(phen)Cl. Highly luminescent rhenium complexes are known, with quantum yields near 0.5 and lifetimes in excess of 10 s. The detection of polarized emission suggests the use of rhenium complexes as probes of the hydrodynamics of large macromolecular complexes and for use in fluorescence polarization immunoassays with gated detection.     

用改进的Van der Pauw法测定方形微区的方块电阻

借助于显微镜放大8×10倍,用目视法将四探针针尖分别控制在样品的面积为100μm×100μm的方形微区的内切圆外四个角区内,利用改进的Van der Pauw法能测定它的方块电阻。测量不受探针游移的影响,无需用测定探针的精确几何位置来进行边缘效应修正。本文利用有限元法对此结论给予证明,并通过金微触点的方块电阻测定得到证实。 关键词：     

Development of calibration techniques for ultrasonic hydrophone probes in the frequency range from 1 to 100 MHz

The primary objective of this work was to develop and optimize the calibration techniques for ultrasonic hydrophone probes used in acoustic field measurements up to 100 MHz. A dependable, 100 MHz calibration method was necessary to examine the behavior of a sub-millimeter spatial resolution fiber optic (FO) sensor and assess the need for such a sensor as an alternative tool for high frequency characterization of ultrasound fields. Also, it was of interest to investigate the feasibility of using FO probes in high intensity fields such as those employed in HIFU (high intensity focused ultrasound) applications. In addition to the development and validation of a novel, 100 MHz calibration technique the innovative elements of this research include implementation and testing of a prototype FO sensor with an active diameter of about 10 μm that exhibits uniform sensitivity over the considered frequency range and does not require any spatial averaging corrections up to about 75 MHz. The results of the calibration measurements are presented and it is shown that the optimized calibration technique allows the sensitivity of the hydrophone probes to be determined as a virtually continuous function of frequency and is also well suited to verify the uniformity of the FO sensor frequency response. As anticipated, the overall uncertainty of the calibration was dependent on frequency and determined to be about ±12% (±1 dB) up to 40 MHz, ±20% (±1.5 dB) from 40 to 60 MHz and ±25% (±2 dB) from 60 to 100 MHz. The outcome of this research indicates that once fully developed and calibrated, the combined acousto-optic system will constitute a universal reference tool in the wide, 100 MHz bandwidth.     

共振光散射技术测定核酸的研究进展

核酸分析是生命科学研究中最重要的技术之一。目前主要是应用核酸内源紫外吸收光谱的紫外分光光度法和基于荧光探针分子与核酸相互作用的荧光分光光度法。紫外分光光度法灵敏度低 ,荧光分光光度法试剂昂贵 ,有毒性。近年来 ,共振光散射技术在核酸分析中的应用得到了迅速的发展。核酸的共振光散射分析方法可以用普通的荧光分光光度计进行测定 ,应用安全、便宜的试剂获得很高的灵敏度。简要介绍了共振光散射分析的基本原理 ,并对近年来利用共振光散射技术分析核酸的研究进行了评述。内容主要包括利用有机染料分子作为核酸的共振光散射探针的分析方法 ;基于阳离子表面活性剂、金属离子及其络合物以及药物与核酸相互作用的分析方法 ;核酸形成大粒子的散射分析方法。     

EPR Features of Spin-Crossover in Tris(<Emphasis Type="Italic">N</Emphasis>,<Emphasis Type="Italic">N</Emphasis>-Dialkyl-Dithiocarbamato)Iron(III)

The temperature dependence of electron paramagnetic resonance (EPR) spectra of a series of dithiocarbamates Fe(RR′dtc)₃ was studied in the temperature range from 5 to 300 K. A small part of solvated complexes serving as spin probes in the EPR-silent matrix enabled the observation of EPR of the Fe(III) ion in the whole temperature range. The spin transition was revealed in the reduction of the integral intensity of the signal from the high-spin complexes and in the non-monotonous change of the line width with temperature decrease due to the effect of the low-spin complexes with short spin–lattice relaxation times. Below ca. 60 K, the ferromagnetic ordering of the magnetic moments in low-spin particles (“domains”) arising at the spin transition was observed.     

Ultrafast-ultrafine probing of high-speed electrical waveforms using a scanning force microscope with photoconductive gating

Picosecond photoconductivity in low-temperature-grown GaAs (LT GaAs) has been used to provide temporal resolution both in rigid probes and in scanning force microscope probes. This article reviews the fabrication and use of such probes. 2.5 ps temporal resolution and few microvolts sensitivity are obtained at arbitrary points on circuits with a spatial definition of 100 nm. Rigid probes are tested in application to analogue and digital circuits. As an alternative to electron beam testing, scanning force probes are applied toin situ imaging and waveform measurement. Finally, the use of time-resolved waveform analysis with scanning-force microscopy probes with semiconductor laser sources is demonstrated.     

镧系配合物的荧光光谱

对镧系配合物荧光光谱的研究进展进行了评述,并对其光物理、光化学进行了归纳、分类,特别是对目前研究非常活跃的镧系离子荧光探针法在配合物和生物分子体系结构探测中的应用情况进行了总结。     

Lanthanide-Sensitized Luminescence as a Promising Tool in Clinical Analysis

Abstract

The recent applications and novelties of lanthanide-sensitized luminescence (LSL) as a detection technique in clinical analysis are here reviewed. In LSL, lanthanide ions form complexes with organic compounds; in these complexes, the energy absorbed by the organic chromophore (usually the analyte) at its characteristic excitation wavelength is transferred to a triplet state of the molecule and then transferred to a resonance level of the lanthanide ion, which finally emits luminescence at its particular emission wavelength. The characteristics of this process will be reviewed and particular attention will be paid to the development of automatic methods of analysis, fluorescence probes, or flow-through optosensors, due to their potential applications in clinical analysis. A critical discussion of the advantages and handicaps of each analytical method is done and the trends of analytical chemistry in this research field are also presented.     

Luminescence-Based Oxygen Sensors: ReL(CO)3Cl and ReL(CO)3CN Complexes on Copolymer Supports

A new class of luminescent rhenium complexes has been tested as oxygen sensors based on luminescent quenching. ReL(CO)₃Cl and ReL(CO)₃CN (L = 2,2-bipyridine or 1,10-phenanthroline and substituted analogues) have several features that seem to indicate suitability as oxygen sensors. These include simple synthesis, long excited-state lifetimes, and high luminescence quantum yields. Intensity and lifetime oxygen quenching measurements were used to study the complexes in various polymer supports including homopolymers of PDMS (polydimethylsiloxane), a methacryloxy containing PDMS (Gp-163), and trimethylsilylmethylmethacrylate (T3642), and copolymers containing Gp-163 and T3642. In contrast to previous studies utilizing [Ru(4,7-diphenyl-1,10-phenanthroline)₃]²⁺ as an oxygen sensor, quenching of the Re complexes proved much more sensitive to the polymer support. With suitable supports, the rhenium chloro complexes demonstrated significant quenching; but the cyano complexes, in spite of being robust in solution, exhibited severe photochemical instability in polymers. The potential of this class of complexes as oxygen sensors and as molecular probes as well as the ramifications in the design of new and different types of sensors is discussed.     

1,10-菲啰啉-四氰基乙烯配合物发光性质的研究

基于探针在近年来得到发展,制备光学性能更为良好的发光材料成为当前化学工作者的研究热点,该工作预制备光学性能更为优良的新型发光材料,以满足人们日常生活及医疗等方面的需求。该工作以四氰基乙烯(TCNE)为第一配体,以1,10-菲啰啉(phen)为第二配体,通过分子间的电荷转移,合成1,10-菲啰啉-四氰基乙烯的电荷转移络合物,并对此反应机理进行初步探索。运用紫外光谱法、荧光光谱法和拉曼光谱法对配合物进行表征及发光性质的研究。比较配合物和配体的紫外吸收峰发现,配合物的吸收均源于配体1,10-菲啰啉的吸收,说明TCNE与Phen形成了稳定的络合物。同时分析荧光光谱,发现配合物的发射峰与配体四氰基乙烯相似,可认为配合物的荧光来自于配体的π—π*电子跃迁。从拉曼图谱中可以看出,在1 000~1 600 cm-1处配合物的拉曼强度比TCNE配体有明显的增强。共振拉曼散射在1 000~1 600 cm-1处振动模式被强耦合,由于分子间的电荷转移使得这些共振拉曼峰被强烈增强。分析结果表明,在一定条件下,1,10-菲啰啉能与四氰基乙烯形成稳定的络合物,且光学性能显著增强。上述研究,合成并研究了1,10-菲啰啉-四氰基乙烯荷移络合物的光学性质,为设计、合成荧光性能良好的配合物提供了实验依据, 并为探索和开发新的核酸探针做出了贡献。     

Fluorescence of Styryl Dyes-DNA Complexes Induced by Single- and Two-Photon Excitation

The series of novel monomer and homodimer styryl dyes based on (p-dimethylaminostyryl) benzothiazolium residues were synthesized and studied as possible fluorescent probes for nucleic acids detection. Spectral-luminescent and spectral-photometric properties of obtained dyes in the unbound state and in DNA presence were studied. Fluorescence emission induced by two-photon excitation of dye-DNA complexes in aqueous buffer solution was registered. Two-photon absorption cross section values of the studied dyes in DNA presence were evaluated.     

Compound formation at Pd(100)/In interfaces

PAC spectroscopy has been used to study the compound formation at the Pd(100)/In interface. Indium films of various thicknesses have been deposited onto Pd(100) single crystals. The¹¹¹In PAC probes are either deposited on the Pd(100) surface before covering it with In or introduced into the In metal before producing the interface. Pd/In compounds with polycrystalline structure of various stoichiometry are identified from the detection of significant electric field gradient tensors after isochronal as well as isothemal annealing. In particular we find an ordered structure at the interface directly after In deposition.     

Study of Polycation-Capped Mn:ZnSe Quantum Dots as a Novel Fluorescent Probe for Living Cells

Transition metal manganese ion (Mn²⁺) doped zinc selenide quantum dots (Mn:ZnSe D-Dots) have been considered as a new material for fluorescent probes in biological labeling. However, this application is limited by the low membrane permeability of D-Dots. In this work, Mn:ZnSe D-Dots were capped with the polycation Sofast to label living cells. For the first time, the efficiency of cellular uptake in living cells is significantly enhanced. Various molar ratios of Sofast to D-Dots were explored and compared to obtain the optimal reaction conditions between Sofast and D-Dots for preparing Sofast/D-Dots nano-compound. A comparison on the fluorescence labeling ability of living cells were made between Sofast/D-Dots and pure D-Dots. Results from laser scanning confocal microscope show that Sofast/D-Dots complexes enter the cells more efficiently than pure D-Dots, even with a lower concentration and shorter incubation time. The cytotoxicities of D-Dots and Sofast/D-Dots were also studied. It was found that Sofast/D-Dots have a much lower cytotoxicity than cadmium-containing quantum dots (i.e. CdTe and CdTe/ZnS). Our results suggest that the non-heavy-metal-containing Sofast/D-Dots complexes have a great potential in the application of biological labeling, especially of long-time bioimaging in living cells.     

Perturbed angular correlation experiments on111In in oxidized AgCd alloy

The hyperfine interaction of¹¹¹Cd probes in preoxidized and α-irradiated Ag_0.94Cd_0.06 alloy has been investigated by PAC measurements and compared with the results for pure Ag. Several probe atom sites were distinguished and characterized by their hyperfine parameters and temperature evolution. The different behavior of In-oxygen and In-oxygen-vacancy complexes in AgCd and Ag was found and discussed. A considerable influence of the irradiation temperature and sample thickness on the different In-oxygen-vacancy complexes formation was evidenced. The parameters of¹¹¹Cd quadrupole interaction measured during isochronal annealing studies in preoxidized and deuteron irradiated AgCd were very similar to the ones observed after α-irradiation.     

Molecular Switching in the Near Infrared (NIR) to Visible/NIR f-f emission with a Functional-Lanthanide Complexes

We report the first observation of three photon induced NIR emission (890-1,400 nm) with NIR excitation (800 nm) from a newly synthesized organic neodymium complex with known molecular structure, suggests the potential use of lanthanide probes for use in three-dimensional imaging and further the study of multi- photon induced emission process in organic-lanthanide complexes.     

On Auger electron spectroscopic measurements of ZnCl2 —coal interaction

By combining electron stimulated desorption (ESD) with low energy electron diffraction (LEED), Auger electron spectroscopy (AES) and work function change (Δφ) measurements the information content of ESD with regard to surface structure and composition is examined, using the surface systems O/W(100) and O/W(110). Although it is not possible to separate the local interaction from the ion escape phase, the comparison of the ESD results with Information derived from LEED, AES and Δφ and the use of simple models of the local interaction gives a rather detailed picture of the location and environment of adsorbed atoms which provides a reasonably reliable basis for the interpretation of UPS spectra of adsorption layers.ESD is extremely sensitive to adsorbed layers. The fact that the ion signal depends not only on coverage but also on the structure and structure-dependent properties of the adsorbate makes on the one hand coverage determination difficult if not impossible, on the other hand opens the door to structure analysis. The potential for obtaining structure information can be easily assessed by comparison with electron probe results.In comparison with other ion probes such as ion scattering spectroscopy and secondary ion mass spectroscopy, ESD is at present the most promising ion probe method for obtaining information on the location of adsorbed atoms from angular and energy distribution measurement (ESDIAD and ESDIED). This is clearly seen by the comparison with the structural data derived from LEED, AES and Δφ measurements for the complex system O/W(100). The consistency of the data obtained with ESD and electron probe techniques lends strong support to the simple models on which the analysis of the ESD results from chemisorbed layers are based. The comparison of ESD results from the system O/W(100) at high coverage and from O/W(110) with 0⁺ ion emission from oxides shows, however, that caution is in place when assigning ESD features to atoms chemisorbed on the metal surface. Without a careful analysis of the ion energy, threshold and/or cross-section such ions cannot be distinguished from ions produced by dissociation of oxides which may be present on the surface only in small quantity. These ions usually are not related to the chemisorbed species which covers most of the surface and therefore dominates the signals seen with (nearly) all other surface probes.If the consistency of LEED, AES, Δφ and ESD data for O/W(100) is not fortuitous, then ESD has already given some important feed-back to the electron probe techniques: the structural models derived from vibrational ELS spectra have to be revised. Increasing accumulation of experimental data and deepening of the theoretical understanding of the physical processes involved in ion emission will have to show how much further information complementary to that from electron probes can be obtained from ion probes.