Fluorescence and Phosphorescence Analysis
01 Feb 1967-Vol. 4, Iss: 5, pp 139-140
TL;DR: Parker et al. as discussed by the authors outline the principles of all these techniques and draw attention to some unusual results obtained with them, including the observation of delayed fluorescence, which was until recently of interest only to the photochemist, but it has now been shown to have potential value as an analytical technique also.
Abstract: From its first analytical applications (Bowman et al, 1955: Parker, et al, 1957) the technique of fluorescence photoelectric spectrometry developed within a decade to become a standard technique for trace organic and inorganic analysis. Reviews and text-books (Bartholomew, 1958: Parker, et al, 1962: Hercules, 1966) deal with the principles of its analytical application and the excellent text book of Udenfriend (1962) covers the applications in the biochemical field. The possibility of using phosphorescence measurement for analytical purposes was first suggested by Lewis and Kasha (1944) and was subsequently taken up by Freed and Salmre (1958) and by Parker and Hatchard (1962). It now promises to rival fluorescence measurement as a method for the trace analysis of organic materials. The observation of delayed fluorescence was until recently of interest only to the photo-chemist, but it has now been shown to have potential value as an analytical technique also (Parker et al, 1965). The object of this paper is to outline briefly the principles of all these techniques and to draw attention to some unusual results obtained with them.
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TL;DR: Flavonoids are plant pigments that are synthesised from phenylalanine, generally display marvelous colors known from flower petals, mostly emit brilliant fluorescence when they are excited by UV light, and are ubiquitous to green plant cells.
2,424 citations
TL;DR: Several, but not all, results support the raft concept, but further definition of the structure, dynamics and function of lipid domains in various biological contexts is urgently required.
412 citations
TL;DR: In this paper, a system of first-order linear differential equations containing as only parameters the rate of fluorescence emission and rate of transport of the excitation from one orthogonal component of the emission to another is described.
Abstract: The depolarization of the fluorescence of solutions by either Brownian rotations or intermolecular energy transfer may be simply described by a system of first‐order linear differential equations containing as only parameters the rate of fluorescence emission and the rate of transport of the excitation from one orthogonal component of the emission to another. The steady‐state solution has the form of Perrin's equation describing the depolarization by Brownian rotations, and the time‐dependent depolarization following a unit light impulse is that originally described by Jablonski. The solution for sinusoidal excitation is novel in that: 1. It shows the difference in lifetime between the polarized components of the emission to be a sensitive function of the ratio of the modulation frequency ω to the emission rate λ. For ω/λ > 1 the difference between the polarized lifetimes may become many times greater than that observed after a unit light impulse. 2. It permits the determination of both the rate of transport of the excitation and the limiting polarization of the fluorescence from observations at one fixed temperature and viscosity. 3. It allows the definition of conditions under which the true or exponential decay of the fluorescence may be measured. Experimental tests of the theory by phase fluorometry are described: These include observations upon dilute solutions in media of limited viscosity where Brownian motion is the only cause of depolarization and observations upon concentrated frozen solutions where depolarization is due to energy transfer alone.
345 citations
TL;DR: Substitution of 5-bromodeoxyuridine for thymidine in synthetic polynucleotides, DNA, or unfixed chromatin quenches the fluorescence of bound dye, and suppression of dye fluorescence permits optical detection of BrdU incorporation associated with DNA synthesis in cytological chromosome preparations.
Abstract: The interaction of the bisbenzimidazole dye 33258 Hoechst with DNA and chromatin is characterized by changes in absorption, fluorescence, and circular dichroism measurements. At low dye/phosphate ratios, dye binding is accompanied by intense fluorescence and circular dichroism and exhibits little sensitivity to ionic strength. At higher dye/phosphate ratios, additional dye binding can be detected by further changes in absorptivity. This secondary binding is suppressed by increasing the ionic strength. A-T rich DNA sequences enhance both dye binding and fluorescence quantum yield, while chromosomal proteins apparently exclude the dye from approximately half of the sites available with DNA. Fluorescence of the free dye is sensitive to pH and, below pH 8, to quenching by iodide ion. Substitution of 5-bromodeoxyuridine (BrdU) for thymidine in synthetic polynucleotides, DNA, or unfixed chromatin quenches the fluorescence of bound dye. This suppression of dye fluorescence permits optical detection of BrdU incorporation associated with DNA synthesis in cytological chromosome preparations. Quenching of 33258 Hoechst fluorescence by BrdU can be abolished by appropriate alterations in solvent conditions, thereby revealing changes in dye fluorescence of microscopic specimens specifically due to BrdU incorporation.
336 citations
TL;DR: A novel fluorescence polarization (FP) method that measures the capacity of a competitor chemical to displace a high affinity fluorescent ligand from purified, recombinant human ER-[alpha] at room temperature shows promise as a high throughput screening method for large-scale testing of environmental and industrial chemicals for ER binding interactions.
Abstract: Over the last few years, an increased awareness of endocrine disrupting chemicals (EDCs) and their potential to affect wildlife and humans has produced a demand for practical screening methods to identify endocrine activity in a wide range of environmental and industrial chemicals. While it is clear that in vivo methods will be required to identify adverse effects produced by these chemicals, in vitro assays can define particular mechanisms of action and have the potential to be employed as rapid and low-cost screens for use in large scale EDC screening programs. Traditional estrogen receptor (ER) binding assays are useful for characterizing a chemical's potential to be an estrogen-acting EDC, but they involve displacement of a radioactive ligand from crude receptor preparations at low temperatures. The usefulness of these assays for realistically determining the ER binding interactions of weakly estrogenic environmental and industrial compounds that have low aqueous solubility is unclear. In this report, we present a novel fluorescence polarization (FP) method that measures the capacity of a competitor chemical to displace a high affinity fluorescent ligand from purified, recombinant human ER-[alpha] at room temperature. The ER-[alpha] binding interactions generated for 15 natural and synthetic compounds were found to be similar to those determined with traditional receptor binding assays. We also discuss the potential to employ this FP technology to binding studies involving ER-ss and other receptors. Thus, the assay introduced in this study is a nonradioactive receptor binding method that shows promise as a high throughput screening method for large-scale testing of environmental and industrial chemicals for ER binding interactions.
311 citations
References
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Book•
01 Jan 1962
TL;DR: Fluorescence assay in biology and medicine is an assay fororescence-based diagnosis of leukaemia and its role in research and treatment is still under investigation.
Abstract: Fluorescence assay in biology and medicine , Fluorescence assay in biology and medicine , مرکز فناوری اطلاعات و اطلاع رسانی کشاورزی
821 citations
TL;DR: In this article, the rotation of 17-isopregnan-3-a-oI-20-one with methanol was reported to be [a]*'D -28.2 * 8' (24.6 * 5'' (C Y ~ ~ D $1.34 * 0.1''; 1 = 2 dm.).
Abstract: hours the rotation was again taken: [ C ~ ] ~ D +77.2 * 5\" (a% +1.75 * 0.1'; 1 = 2 dm.). Treatment of 17-Isopregnan-3( a)-ol-20-one with Hydrogen chloride.-In a manner similar to the above a solution of 44.7 mg. of 17-isopregnan-3(a)-ol-20-one made up to 5 ml. with 4.3% methanolic hydrogen chloride was refluxed for four hours and then the rotation was taken: [ a I s 1 ~ $75.6 * 5\" ( C Y ~ ~ D $1.35 * 0.1\"; 1 = 2 dm.). Acetate of 17-bopregnan-3( ~~)-ol-2O-one.-A solution of 100 mg. of 17-isopregnan-3(a)-oI-20-one in 1 ml. of acetic anhydride and 0.66 ml. of acetic acid was refluxed for twenty minutes. After standing for three hours the solution was diluted to turbidity with water. The acetate separated in fine needles and was collected, weight 95 mg. After one recrystallization from methanol it melted a t 157-159' and mixed melting points with b:th the normal and 17-isopregnanolones gave about 30 depressions: [a]*'D -28.2 * 8' (24.16 mg. made up to 2 cc. with methanol; 1 = 1 dm.; a a 4 ~ -0.34 * 0.1'). Anal. Calcd. for CzzH380~: C, 76.61; H, 10.07. Found: C, 76.82; H, 10.32. Oxime of 17-Isopregnan-3(a)-oI-20-one.-A solution of 100 mg. of 17-isopregnan-3(a)-oI-20-one, 100 mg. of hydroxylammonium chloride, and 150 mg. of sodium acetate (trihydrate) in 2 ml. of methanol and 0.5 d. of water was refluxed for two hours. After cooling the solution was poured into water, extracted with ether, and the ether solution was dried over anhydrous sodium sulfate. The ether was removed and the residue was crystallized first from dilute acetone and then from methanol giving a white crystalline oxime melting at 192-197\" (with decomposition). A mixed melting point with a sample of the oxime of the normal pregnan-3(~~)-01-20-one@ gave a depression of about 20\". Anal. Calcd. for ColHssOzN: N, 4.20. Found: N, 4.57. Test with Digitonin.-To about 1 mg. of 17-isopregnan3(a)-ol-20-one in a few drops of 90% methanol was added an equal volume of a saturated solution of digitonin in 90% methanol. No precipitate was obtained even after standing for several hours.
627 citations
"Fluorescence and Phosphorescence An..." refers methods in this paper
...The possibility of using phosphorescence measurement for analytical purposes was first suggested by Lewis and Kasha (1944) and was subsequently taken up by Freed and Salmre (1958) and by Parker and Hatchard (1962)....
[...]
TL;DR: This volume is a remarkably complete summary of a rapidly advancing field of fluorescent measurement, with methods for assay of antibiotics such as griseofulvin and the tetracyclines, catecholamines, plus fluorescent microscopy and specific applications.
Abstract: Methods for quantitative fluorescent analysis reemphasize the dependence of scientific advances on technologic development. The techniques of fluorescent assay are versatile and relatively simple. Their sensitivity is comparable to that of radioactive assay methods, making possible the detection of trace amounts of many substances. This volume is a remarkably complete summary of a rapidly advancing field. A refreshing introduction reviews the relationship of physicists and biologists to the development of techniques for fluorescent assay and is followed by an essentially nonmathematical treatment of luminescence and fluorescence. Sections on instrumentation are detailed. The ultrasensitivity of fluorescence analysis leads to problems of contamination, self-absorption, and quantitative recovery similar to those with radioactive isotopes. These problems are carefully delineated by the author. Specific applications of fluorescent measurement constitute the bulk of the book. Of particular interest are methods for assay of antibiotics such as griseofulvin and the tetracyclines, catecholamines, plus fluorescent microscopy and
188 citations
"Fluorescence and Phosphorescence An..." refers background or methods in this paper
...Reviews and text-books (Bartholomew, 1958: Parker, et al, 1962: Hercules, 1966) deal with the principles of its analytical application and the excellent text book of Udenfriend (1962) covers the applications in the biochemical field. The possibility of using phosphorescence measurement for analytical purposes was first suggested by Lewis and Kasha (1944) and was subsequently taken up by Freed and Salmre (1958) and by Parker and Hatchard (1962). It now promises to rival fluorescence measurement as a method for the trace analysis of organic materials....
[...]
...Reviews and text-books (Bartholomew, 1958: Parker, et al, 1962: Hercules, 1966) deal with the principles of its analytical application and the excellent text book of Udenfriend (1962) covers the applications in the biochemical field....
[...]
...Reviews and text-books (Bartholomew, 1958: Parker, et al, 1962: Hercules, 1966) deal with the principles of its analytical application and the excellent text book of Udenfriend (1962) covers the applications in the biochemical field. The possibility of using phosphorescence measurement for analytical purposes was first suggested by Lewis and Kasha (1944) and was subsequently taken up by Freed and Salmre (1958) and by Parker and Hatchard (1962)....
[...]
188 citations