Comparisons between the luminol light standards and a new method for absolute calibrations of light detectors

Michael, Paul R.; Faulkner, Larry R.

doi:10.1021/ac50002a031

Cited by 21 publications

(12 citation statements)

References 22 publications

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“…However, when H 2 O 2 was further increased to 5 × 10 −3 M , the quantum yield decreased to 1.0(±0.17)%. As discussed below, the optimized quantum yield of 1.23(±0.20)% in the present experiment shows very good agreement with the value of 1.24% determined by the majority of previous related reports (9–18). We do not go into more detailed H 2 O 2 ‐concentration dependence or microscopic chemical mechanisms for the dependence, which is beyond the purpose of this paper.…”

Section: Resultssupporting

confidence: 92%

“…Quantum yields of popular bio/chemiluminescence reaction systems such as firefly (1), cypridina (2), aequorin (3–5), bacteria (6–8) and luminol (9) were mostly reported in 1960–1970. Luminol chemiluminescence was later reexamined by other elaborate methods (10–18) and is now often used as one of the secondary light standards (17–19) in quantitative luminescence measurements. In each of these experiments, however, the absolute sensitivity of each detector had to be calibrated with a standard of light.…”

Section: Introductionmentioning

confidence: 99%

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Development of a Quantitative Bio/Chemiluminescence Spectrometer Determining Quantum Yields: Re‐examination of the Aqueous Luminol Chemiluminescence Standard

Ando

Niwa

Yamada

et al. 2007

Photochem & Photobiology

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We have developed a bio/chemiluminescence spectrometer with a cooled charge-coupled-device (CCD) detector to obtain a quantitative luminescence spectrum as the absolute number of all emitted photons at each wavelength. The integrated area of the spectrum divided by the number of reacted substrate molecules gives the quantum yield. Calibration of the absolute sensitivity of the CCD-spectrometer system was performed by using lasers and a tungsten lamp with calibrated powers as primary light standards, and calibration of the light-collection efficiency of the spectrometer with several kinds of cells for liquid samples was achieved by introducing a simple reference double-plate cell. The reference cell is not convenient for final bio/chemiluminescence measurements but is useful for the calibration because it has well-defined angular dependence of light emission, allowing accurate calculation of the light-collection efficiency. Using this CCD-spectrometer system, we re-examined the quantum yield of aqueous luminol chemiluminescence with H2O2 catalyzed by horseradish peroxidase. The quantum yield was constant for a wide range of luminol concentrations, whereas it changed and had an optimum against H2O2 concentrations. The optimum quantum yield was 1.23(+/-0.20)%, which is in good agreement with previously reported values.

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Section: Resultssupporting

confidence: 92%

Section: Introductionmentioning

confidence: 99%

Development of a Quantitative Bio/Chemiluminescence Spectrometer Determining Quantum Yields: Re‐examination of the Aqueous Luminol Chemiluminescence Standard

Ando

Niwa

Yamada

et al. 2007

Photochem & Photobiology

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“…The blue fluorescence protein was routinely assayed by its fluorescence intensity at 470 nm when excited at 420 nm, with an Aminco-Bowman Spectrofluorimeter. Luciferase activity was determined with a digital photometer, designed and constructed by G. J. Faini, which was calibrated for absolute photon sensitivity with the luminol chemiluminescence reaction as a light standard (22); This standard is directly traceable to the National Bureau of Standards (NBS) Lamp (23) and its calibration has been confirmed by three independent methods (24)(25)(26). NADH dehydrogenase was purified from P. fischeri 399 by C. White.and coupled efficiently with all types of luciferases used.…”

Section: Methodsmentioning

confidence: 99%

Isolation of the in vivo emitter in bacterial bioluminescence

Gast

Lee

1978

Proc. Natl. Acad. Sci. U.S.A.

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A blue fluorescence protein has been isolated and purified from extracts of the luminous bacterium Photobacterium phosphoreum. It is a single polypeptide of molecular weight 22,000 with absorption maxima at 274 and 418 nm. It is efficiently fluorescent (OF 0.45), with a fully corrected spectral maximum (476 nm) and distribution identical to the in vivo bioluminescence from this same type of bacterium. At low concentration this fluorescence shifts towards the red and becomes identical to the in vitro bioluminescence emission. This spectral shift apparently results from a change in the protein pulled by dissociation of the chromophore . If the blue fluorescence protein is included in the in vitro bioluminescence reaction with reduced FMN, oxygen, aldehyde, and luciferase (P. phosphoreum), the bioluminescence spectrum is shifted towards the blue from its maximum at 490 nm to one at 476 nm, where it is again identical in all respects to the in vivo bioluminescence spectrum. This is accompanied by an increase in the initial light intensity by an order of magitude at saturating levels of blue fluorescence protein, and the specific light yield of the luciferase is increased 4-fold. It is suggested that the blue fluorescence protein acts as a sensitizer of the bacterial bioluminescence reaction. (14) claimed that the emitter must be some sort of flavin-derived species (2), three proposals for its structure were put forward, supported almost solely by the similarity of the fluorescence of model compounds to the bioluminescence spectra. Eley et al. (12) (17) proposed an FMNH2 molecule substituted in the 4a-position.It had been tacitly assumed that the mechanism of reaction and identity of the emitter are the same in vivo as in vitro. The possibility of a difference has been raised by the recent discovery of a bacterial type emitting at 545 nm (18). In this paper we show that a protein-bound chromophore can be isolated from extracts of the bioluminescent bacteria P. phosphoreum that is closely associated with luciferase and that fulfills all the conditions to qualify it as the in vivo emitter. MATERIALS AND METHODSThe bacterium Beneckea harveyi, strain 392 in the classification scheme of Reichelt and Baumann (19), previously designated "MAV,"' was obtained from J. W. Hastings (Harvard University). The type "A-13" was isolated from the light organ of the "silver macrourid" fish by J. Paxton (Australian Museum) and has been identified as Photobacterium phosphoreum (J. Fitzgerald, private communication). The type Photobacterium fischeri, strain 399, was obtained from F. H. Johnson (Princeton University). The bacteria were grown and the luciferase and FMN were purified as described (20,21). The blue fluorescence protein was routinely assayed by its fluorescence intensity at 470 nm when excited at 420 nm, with an Aminco-Bowman Spectrofluorimeter. Luciferase activity was determined with a digital photometer, designed and constructed by G. J. Faini, which was calibrated for absolute photon sensitivity with the luminol ch...

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“…Thus +cL for the luminol oxidation has been measured for radiometry using a light source comprised of a mixture of scintillators in a 14C labelled solvent (Hastings and Weber, 1963) and several spectrometric methods including calibrated spectroradiometers and photomultipliers (Lee and Seliger, 1972;Roberts and Hirt, 1967;Rauhut et al, 1966), matching of corrected and uncorrected spectra with that of a standard lamp viewed through filter combinations (band emission approximation method) (Lee and Seliger, 1965), and an integrating sphere detector with a quantum counter (Bezman and Faulkner, 1971;Michael and Faulkner, 1976).…”

Section: Introductionmentioning

confidence: 99%

“…Although the quantum yields of many chemiluminescent reactions have been measured (Mellor and Richter, 1974) the oxidation of luminol appears to be the only reaction that has been widely used and tested (Hastings and Weber, 1963;Seliger, 1965, 1972;Roberts and Hirt, 1967;Rauhut et al, 1966;Bezman and Faulkner, 1971). Thus +cL for the luminol oxidation has been measured for radiometry using a light source comprised of a mixture of scintillators in a 14C labelled solvent (Hastings and Weber, 1963) and several spectrometric methods including calibrated spectroradiometers and photomultipliers (Lee and Seliger, 1972;Roberts and Hirt, 1967;Rauhut et al, 1966), matching of corrected and uncorrected spectra with that of a standard lamp viewed through filter combinations (band emission approximation method) (Lee and Seliger, 1965), and an integrating sphere detector with a quantum counter (Bezman and Faulkner, 1971;Michael and Faulkner, 1976).…”

Section: Introductionmentioning

confidence: 99%

Determination of absolute chemiluminescence quantum yields for reactions of bis‐(pentachlorophenyl) oxalate, hydrogen peroxide and fluorescent compounds

Catherall

Palmer

Cundall

1989

J. Biolumin. Chemilumin.

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Absolute chemiluminescence quantum yields (phi CL) for reactions of bis-(pentachlorophenyl) oxalate (PCPO), hydrogen peroxide (H2O2) and 9:10 diphenyl anthracene (DPA) have been determined. A fully corrected chemiluminescence monitoring spectrometer was calibrated for spectral sensitivity using the chemiluminescence of the bis-(pentachlorophenyl) oxalate system as a liquid light source, the total photon output of which had previously been determined by chemical actinometry. At high (PCPO)/(H2O2) ratios phi CL was found to be independent of PCPO and H2O2 concentrations.

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Comparisons between the luminol light standards and a new method for absolute calibrations of light detectors

Cited by 21 publications

References 22 publications

Development of a Quantitative Bio/Chemiluminescence Spectrometer Determining Quantum Yields: Re‐examination of the Aqueous Luminol Chemiluminescence Standard

Development of a Quantitative Bio/Chemiluminescence Spectrometer Determining Quantum Yields: Re‐examination of the Aqueous Luminol Chemiluminescence Standard

Isolation of the in vivo emitter in bacterial bioluminescence

Determination of absolute chemiluminescence quantum yields for reactions of bis‐(pentachlorophenyl) oxalate, hydrogen peroxide and fluorescent compounds

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