Analytical techniques, very much like women's clothing styles, tend to go in and out of vogue. This, however, is where the similarity ends. Whereas women's styles are the result of the whims of the designers, analytical methods remain popular only as long as they can perform a particular measurement better, easier, and/or cheaper than a competitive approach. The ability of a technology to survive is thus dependent on state-of-the-art instrumentation and analytical procedures but is restrained by the inherent limitations of the method.
Picosecond fluorescence spectroscopy was used to investigate the ultrafast energy transfers occurring in the light-harvesting accessory pigment complex of both photosynthetic and nonphotosynthetic leaves of the Norway maple (acer platenoides). The time-resolved fluorescence emission was measured for various wavelength regions both above and below 600 nm at room temperature and 90 K. Excitation was provided by single 6 ps pulses at 530 nm obtained from the frequency-doubled output of a Nd:glass laser, and the fluorescence emission was analyzed with the use of a streak camera optical multichannel analyzer system. Nearly identical decay kinetics were observed in both the nonphotosynthetic and photosynthetic systems. These results highlight the close relationship between the physical configuration of the carotenoid and chlorophyll accessory pigment molecules on the light-harvesting lattice and their energy transfer properties. A trapping model incorporating both photoactive and non-photoactive traps as quenchers of the absorbed excitation energy is proposed. These results question the interpretation of the in vivo fluorescence decay kinetics as being indicative of the photoactive state of the light-harvesting complex.
The measurement of the lifetimes of excited states of molecules by picosecond time-resolved fluorescence methods is discussed. Attention is focused on the methods that involve the use of mode-locked solid-state and dye lasers as excitation sources. Time-correlated photon counting and direct recording using streak cameras are the diagnostic methods that have proved most useful at times of less than 1 ns. The second section of the report outlines some work carried out at the Center for Fast Kinetics Research in which the variation of fluorescence lifetime of some xanthene dyes with changes in their environment has been used to deduce information on the strengths of hydrogen bonds therein. Both neat organic solvents and aqueous surfactant micelles have been employed.
The use of remote analysis techniques over optical fibers for underground water sampling is considerably less expensive (and offers faster and more complete analysis) than the conventional borehole fields containing sampling devices. The technology is based on fluorometric analysis, combining long range communication fiber optics, laser excitation, and Raman spectroscopic measurement technology. Detection at up to 1000 m range of ppb quantities of fluorescent dye tracers, and ppm quantities of heat and radiation stable tracers, with ~1% qualitative accuracy, has been accomplished. For nonfluorescent compounds, or for mixtures too complex for the intrinsic selectivity of fluorescence, specific compound optrodes (by analogy to electrodes) are used at the fiber tip. These contain immobilized reagents which become fluorescent or whose fluorescence is altered by the sample (reversibly for continuous measurements or irreversibly for cumulative ones).
A detailed description is given of a recently established life-times method, in which the time decay of atomic fluorescence is recorded following pulsed laser excitation of an atomic vapor produced by cathodic sputtering in a rare-gas discharge. The method is readily applicable to neutral and singly ionized atoms of a wide range of elements, including the highly refractory elements for which, until very recently, little lifetime data have been available. A theoretical discussion is presented of the form of the time-resolved fluorescence signal detected following pulsed laser excitation of a group of atoms, with particular emphasis on possible sources of distortion in the fluorescence decay curves. Ex-amples are given of lifetime measurements on both neutral and singly ionized atoms of a number of refractory elements and comparisons made with other recent laser-induced fluorescence measurements. Some new lifetime data are reported for levels in Zr I, Zr II, Ag I, and Cu I.
Laser-induced fluorescence can be used to detect with high sensitivity small molecular species, typically free radicals, which are the intermediates in the chemistry of combustion processes, the atmosphere, and plasmas. Using as examples recent work from our laboratory, we describe the laser spectroscopy and spectroscopically based collision studies needed for application of the laser-induced fluorescence techniques, with an emphasis on combustion diagnostics.
Molecular multiphoton excitation with high power ultraviolet lasers allows access to highly excited states which may undergo fragmentation to produce a variety of interesting excited-state products. Recent work on photo-fragment fluorescence following excimer laser photolysis of various small molecules is reviewed, and aspects related to the mechanisms of excitation and fragmentation and various practical implications are discussed.
The use of laser-induced matrix-isolation fluorescence spectrometry for identification and quantification of individual fluorescent constituents of complex samples is surveyed. The advantages of matrix isolation are outlined, and the advantages of laser excitation in low-temperature fluorometry are discussed. The use of Shpol'skii matrices, optical site-selection techniques, and time resolution in matrix-isolation fluorometric analyses is considered. The relationship between selectivity and analytical detection limits in the fluorometric analysis of complex samples is emphasized.
Recent advances in excitation methods of molecules in the gas phase and in detection and data acquisition techniques are utilized for studying the photophysics of N2. Selective excitation of high lying electronic states of the molecule is achieved by applying a pulsed electrical discharge followed by a pulsed laser beam. By monitoring and analyzing the wavelength- and time-resolved fluorescence, new information on fast collisional coupling between the electronic states is obtained. This information is crucial to the understanding of photophysical and photochemical processes in systems containing excited nitrogen, including atmospheric phenomena such as auroras and airglows.
Due to its sensitivity and selectivity, luminescence detection is often the choice in a wide variety of analytical applications. However, until recently only one parameter of luminescence was being monitored in a single experiment, for example, emission spectra scanned at a fixed single excitation wavelength, and decay of luminescence observed at a single wavelength of excitation and emission. In this paper we will present some of the recent developments that provide multiparametric detection of luminescence phenomena, for example, a complete intensity map of phosphorescence, simultaneously, at multiple wavelengths of excitation and emission over time. Another area involves the use of polarized radiation in effecting luminescence detection. We will discuss the instrumentation and data analysis for these kinds of experiments.
Fluorescence excitation spectra of chlorophyll a and flavin, corrected either by employing the common quantum counter rhodamine B or by comparison with the corresponding absorption spectra, are discussed.absorption spectra are not necessarily identical, as commonly assumed. functions, "kernels," is used for smoothing. Cross-correlations with two other, more complex kernels demonstrate their equivalence with the first and fourth derivative. The latter procedures, in turn, have been known for some time to improve wavelength resolution. For the determination of fluorescence quantum efficiencies, spectra are converted from wavelength to wave number on the x-axis, and from energy to quanta absorption on the y-axis. Based on thermodynamic arguments, the fluorescence emission spectrum of chlorophyll a is calculated from its absorption spectrum. Measured and calculated emission spectra are practically identical. Similarly, the integrated absorption spectrum of chlorophyll a was used to calculate the intrinsic fluorescence lifetime and, taking the quantum efficiency of fluorescence into account, the ap-parent lifetime. Continuous fluorescence polarization spectra have been obtained for various commonly used fluorophores. Based on a well-known formula originally deduced by Perrin, "angle spectra" are calculated from these polarization spectra, which immediately exhibit the angles between the electronic transition dipoles. In addition, for chlorophyll a and flavin, "anisotropy spectra" have been determined. Taking the dependency of fluorescence polarization on temperature or on the concentration of the fluorophore into account, both its rotational relaxation time and the value of Rc are determined. For all thirteen fluorophores investigated, the corrected and normalized fluorescence excitation and emission spectra are given, including the apparent 0-0 transition energies as deduced from the wavelength of their intersection.
In this paper several level-crossing detection techniques based on photon statistics are presented. Results on level-crossing detection through the measurement of the normalized second-order factorial moment n(2)(T) are reviewed. Detection by measurement of the normalized intensity correlation function g(2)(r) or the Laplace transform of time-interval probability is studied. The interest of the above mentioned techniques is analyzed.
Recent advances are described in the combined use of fluorometric derivatization and high performance liquid chromatography (HPLC) for clinical chemistry determinations. Derivatives (especially dansyl derivatives) can be formed prior to chromatography in the case of estrogens, amino acids, and catecholamines. In post-column reactions, we preferred to use air-segmented reactions as they conform better to all the optimized chromatographic and spectrofluorometric parameters. Fluorescent derivatives produced from cate-cholamines, tryptophan and its metabolites, hydroxyindoles, tryptamine, amino acids, sugars, polyamines, and other substances are often sufficiently sensitive to be detected in picogram quantities by HPLC. Their reaction principle and some of their applications to samples are described. Recently, chemical excitation of fluorophore-like dansyl amino acid was proposed as a detection system for HPLC. By a post-column reaction, a fluorophore can be made to emit light by its reaction with trichlorophenyl oxalate (TCPO) and hydrogen peroxide. The detection limit of this system is about 10 fmol for each dansyl amino acid. Application of this new reaction to catecholamines opens up new prospects for fluorometric detection.
Laser fluorescence spectroscopy of biomolecules, especially of nucleic acids and DNA-dye-complexes, is reviewed. Fluorescence measurements, energy transfer processes, and selective laser action are discussed.
This review will deal with characteristics of fluorescence analysis in comparison with some other spectroscopic analyses, some thoughts and suggestions on equipment and techniques, and some applications in proteins.
A new spectrophotometer to measure a wide variety of the optical properties of materials, such as unidirectional/bidirectional absolute reflectance and transmittance as a function of (a) wavelength, in the range of 0.3 to 2.0 um with a resolution of 0.005 um, (b) angle of incidence in the range of 3° to 65° with an accuracy of 0.5°,(c)state of polarization, 0°, 45°, and 90° of the incident light, has been designed and developed. It is demonstrated that these optical properties can be measured to an accuracy better than 1%. In addition, this multifunctional instrument is tailored to monitor the relative spectral signatures of living vegetation that has been grown under controlled conditions. These spectral signatures are useful for the effective interpretation of remotely sensed data obtained from aircraft and satellite borne instrumentation.
The design and performance of an 800 X 800 pixel charge-coupled device (CCD) imager are described. This device is fabricated utilizing a three-phase, three-level polysilicon gate process. The chip is thinned to 8 um and is employed in the rear illumination mode. Detailed measurements of the device performance, including dark current as a function of temperature, linearity, and noise, are presented. The device is coated with an ultraviolet (UV) downconverting phosphor which allows imaging with the same device over an extremely wide optical bandwidth.
The present paper considers the aspect of the Michelson interferometer when the beam divider and the compensating plate are not identical parallel plates of glass, but are slightly wedged. The results are utilized to set up tolerances on the wedge angles of these basic components of the interferometer. These tolerances are good enough for the instruments that are used in the teaching laboratory or even for research.
A new hybrid, direct, real-time, and parallel analog-to-digital (A/D) conversion system has been theoretically and experimentally investigated. Basically this system consists of an A/D conversion screen, a photosensor panel, a comparator, and a coder. For the purpose of designing an A/D conversion screen, various quantizer characteristics and the minimization of the mean-square distortion are briefly described. Based on the quantizer characteristics, the design algorithms of the A/D conversion screen are developed. It is found that charge-coupled device (CCD), charge-injection device (CID), and metal oxide semiconductor (MOS) photodiode image sensor arrays can be used for the large-area hybrid A/D conversion system, and photodiode or photocell arrays can be applied for a single-pixel A/D conversion system. An error analysis shows that the conversion errors result from the discrepancy between the fabricated and designed transmittance function of the screen cells and the nonuniform characteristics of the photosensors. Also, this system is theoretically compared with existing electronic A/D conver-sion systems. To show the feasibility of this system, a single-pixel 16-level A/D conversion system using a single-cell 16-level A/D conver-sion screen, a photosensor panel of 16 photocells, and accessory elec-tronic circuits is designed, constructed, and experimented with input op-tical signals. The experimental results proved that this system can be used for achieving A/D conversion of both coherent or incoherent and monochromatic or polychromatic optical signals. The system can be potentially applied for the interface at the optical input of a digital computer for real-time optical data processing, picture coding, and optical telemetry.
Inorganic phosphor screens that are normally designed for low-energy x-ray imaging applications are shown to be useful for imaging with photons and neutrons having MeV energies. Characterizations of a number of commercially available phosphors with regard to detection efficiency for radiation type and energy, effective spatial resolution, optical self-absorption, response time, and spectral emissions are tabulated. Applications to high-energy x-ray imaging, nuclear fuel rod imaging, and nuclear fusion source imaging are shown to be feasible.
A method is described for determining the diameter of a Gaussian beam by measuring the light reflected as the beam scans an "opaque Ronchi ruling" consisting of dark bars on a light background. The effects of unequal bar/space widths are included.
Particle size and velocity measurements have been obtained in a low-speed (6 to 10 m/s), 2800 K combustor 30 cm in diameter. The measurements were obtained using a particle-sizing interferometer coupled to a 0.5 m spectrometer for background light rejection from radiant particles. Results obtained for the combustion of powdered coke clearly indicate the capabilities of this type of instrument to estimate combustor efficiency as a function of temperature. Comparison of the optically sampled measurements with other sampling techniques shows reasonable agreement.
We have made crosstalk measurements on our U-groove isolated photodiode array by two independent techniques, an electrical measurement method and a laser scanning method. These two methods are shown to corre-late with each other. The crosstalk between two adjacnt pixels of the U-groove isolated array was below 5 X 10-4, which compared favorably to the value of 10-1 for the conventional diffusion junction photodiode array. Also, the effective diffusion length of the photogenerated carriers was measured to be 10µm in the N-region and 6 um in the top P-region. The advantages of the detector isolation technique for optical signal processing are presented.
An application of real-time holography to detect microcracks in ancient golden paintings is presented. The technique does not require sophisticated laboratory equipment and enables us not only to pinpoint the presence of a crack in embryo, inaccessible by more conventional methods of inspection, but also to monitor its growth in real time, giving, therefore, a valid aid to artworkers for determining the best conditions for the preservation of paintings.
From the information generated from the above expressions, the temperature gradient across the stagnation boundary and the temperature gradient across the faceplate may be determined. Thus, inside wall temperature will be found from
At an impressive ceremony in Geneva, Switzerland, this April, SPIE inaugurated a major award and prize for out-standing achievement in optics in honor of one of the most distinguished scientist-inventors in the history of our field, Professor Dennis Gabor. Professor Gabor's notable achieve-