Fluorescence excitation source

The photochemical reaction was initiated with a UV source, and an argon-ion laser was used as the fluorescence excitation source (2 = 457.9 nm). A... 

This photochemical deactivation reduces the probability of fluorescence. Thus, this process has to be considered in competition using high-intensity fluorescence excitation sources [7]. For this reason one has to take care in any fluorescence measurement, especially examining photochemically instable compounds. Repetitive quantitative examination of the same sample yields a decreasing fluorescence signal. 

The vast majority of single-molecule optical experiments employ one-photon excited spontaneous fluorescence as the spectroscopic observable because of its relative simplicity and inlierently high sensitivity. Many molecules fluoresce with quantum yields near unity, and spontaneous fluorescence lifetimes for chromophores with large oscillator strengths are a few nanoseconds, implying that with a sufficiently intense excitation source a single... 

The intensity of fluorescence. If, is proportional to the amount of the radiation from the excitation source that is absorbed and the quantum yield for fluorescence... 

The intensity of fluorescence therefore, increases with an increase in quantum efficiency, incident power of the excitation source, and the molar absorptivity and concentration of the fluorescing species. 

Standardizing the Method Equations 10.32 and 10.33 show that the intensity of fluorescent or phosphorescent emission is proportional to the concentration of the photoluminescent species, provided that the absorbance of radiation from the excitation source (A = ebC) is less than approximately 0.01. Quantitative methods are usually standardized using a set of external standards. Calibration curves are linear over as much as four to six orders of magnitude for fluorescence and two to four orders of magnitude for phosphorescence. Calibration curves become nonlinear for high concentrations of the photoluminescent species at which the intensity of emission is given by equation 10.31. Nonlinearity also may be observed at low concentrations due to the presence of fluorescent or phosphorescent contaminants. As discussed earlier, the quantum efficiency for emission is sensitive to temperature and sample matrix, both of which must be controlled if external standards are to be used. In addition, emission intensity depends on the molar absorptivity of the photoluminescent species, which is sensitive to the sample matrix. 

Precision When the analyte s concentration is well above the detection limit, the relative standard deviation for fluorescence is usually 0.5-2%. The limiting instrumental factor affecting precision is the stability of the excitation source. The precision for phosphorescence is often limited by reproducibility in preparing samples for analysis, with relative standard deviations of 5-10% being common. 

Sensitivity From equations 10.32 and 10.33 we can see that the sensitivity of a fluorescent or phosphorescent method is influenced by a number of parameters. The importance of quantum yield and the effect of temperature and solution composition on f and p already have been considered. Besides quantum yield, the sensitivity of an analysis can be improved by using an excitation source that has a greater... 

Better detection limits are obtained using fluorescence, particularly when using a laser as an excitation source. When using fluorescence detection, a small portion of the capillary s protective coating is removed and the laser beam is focused on the inner portion of the capillary tubing. Emission is measured at an angle of 90° to the laser. Because the laser provides an intense source of radiation that can be focused to a narrow spot, detection limits are as low as 10 M. 

Luminescent Pigments. Luminescence is the abihty of matter to emit light after it absorbs energy (see Luminescent materials). Materials that have luminescent properties are known as phosphors, or luminescent pigments. If the light emission ceases shortly after the excitation source is removed (<10 s), the process is fluorescence. The process with longer decay times is referred to as phosphorescence. 

The analysis was performed by XRF method with SR. SRXRF is an instrumental, multielemental, non-destructive analytical method using synchrotron radiation as primary excitation source. The fluorescence radiation was measured on the XRF beam-line of VEPP-3 (E=2 GeV, 1=100 mA), Institute of Nuclear Physics, Novosibirsk, Russia. For quality control were used international reference standards. 

Figure 6. Tempcraiure dependence of the fluorescence lifetime of BMPC in 1 1 ethanol-mcihanol. Measurements were carried out at the LENS laboratory of Florence by a picosecond apparatus using as an excitation source (at 380 nm) a dye laser pumped by a frequency-doubled cw Nd-YAG laser and recording the fluorescence time jirofiles by a streak camera. Since the overall insuumental response time was 75-80 ps, decays with t>200 ps, observed at T<130 K, were analyzed without deconvolution. At 177, 178 and 193 K, the lifetimes were roughly estimated as i=(FWHM -77 ), where FWHM was the width at half maximum of the decay. Because of the rather high sample absorbances (An,x=2), self absorption may have reduced the lifetimes to some extent.

Visible lasers are typically used for sample excitation, although near-IR lasers can be used when visible excitation sources cause sample fluorescence, obscuring the Raman scatter. 

For the investigation of triplet state properties a laser flash photolysis apparatus was used. The excitation source was a Lambda Physik 1 M 50A nitrogen laser which furnished pulses of 3.5 ns half-width and 2 mJ energy. The fluorescence decay times were measured with the phase fluorimeter developed by Hauser et al. (11). 

At present, two main streams of techniques exist for the measurement of fluorescence lifetimes, time domain based methods, and frequency domain methods. In the frequency domain, the fluorescence lifetime is derived from the phase shift and demodulation of the fluorescent light with respect to the phase and the modulation depth of a modulated excitation source. Measurements in the time domain are generally performed by recording the fluorescence intensity decay after exciting the specimen with a short excitation pulse. 

Grant, D. M., Elson, D. S., Schimpf, D., Dunsby, C., Requejo-Isidro, J., Auksorius, E., Munro, I., Neil, M. A. A., French, P. M. W. Nye, E., Stamp, G. and Courtney, P. (2005). Optically sectioned fluorescence lifetime imaging using a Nipkow disk microscope and a tunable ultrafast continuum excitation source. Opt. Lett. 30, 3353-5. 

Now, we are not particular experts in X-ray and gamma-ray spectroscopy (nor mass spectroscopy, for that matter), but our understanding of those technologies is that they are used mainly in emission mode. Even when the exciting source is a continuum source, such as is found when an X-ray tube is used to produce the exciting X-rays for an X-ray Fluorescence (XRF) measurement, the measurement itself consists of counting the X-rays emitted from the sample after the sample absorbs an X-ray from the source. These measurements are themselves the equivalent of single-beam measurements and will thus also be Poisson-distributed in accordance with the basic physics of the phenomenon. 

In conventional chip experiments, fluorescence scanners are used for chip read-out. In the case of laser scanners, HeNe lasers are used as excitation sources and photomultiplier tubes as detectors, whereas CCD-based scanners use white light sources. The optical system can be confocal or non-confocal. Standard biochip experiments are performed using two fluorescent labels as... 

The emission spectra of BODIPY derivatives normally display narrow bandwidths, providing intensely fluorescent labels for biomolecules. Unfortunately, they also have very small Stoke s shifts, typically on the order of only 10-20 nm. Excitation at the optimal wavelength may cause some interference in measurements at the emission wavelength due to light scattering or cross-over from the wide bandwidth of the excitation source. The dyes usually require excitation at sub-optimal wavelengths to prevent this problem. 

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