Secondary fluorescence

Ema data can be quantitated to provide elemental concentrations, but several corrections are necessary to account for matrix effects adequately. One weU-known method for matrix correction is the 2af method (7,31). This approach is based on calculated corrections for major matrix-dependent effects which alter the intensity of x-rays observed at a particular energy after being emitted from the corresponding atoms. The 2af method corrects for differences between elements in electron stopping power and backscattering (the correction), self-absorption of x-rays by the matrix (the a correction), and the excitation of x-rays from one element by x-rays emitted from a different element, or in other words, secondary fluorescence (the f correction). 

In another approach, which was previously mentioned, the mass thickness, or depth distribution of characteristic X-ray generation and the subsequent absorption are calculated using models developed from experimental data into a < )(p2) function. Secondary fluorescence is corrected using the same i flictors as in ZAP. The (pz) formulation is very flexible and allows for multiple boundary conditions to be included easily. It has been used successfully in the study of thin films on substrates and for multilayer thin films. 

A consequence of absorption of X rays is the inner shell ionization of the absorbing atoms and the subsequent generation of characteristic X rays from the absorbing atoms, called secondary fluorescence, which raises the generated intensity over that produced by the direct action of the beam electrons. Secondary fluorescence can be induced by both characteristic and bremsstrahlung X rays. Both effects are compo-sitionally dependent. 

Although XRF is generally the X-ray spectrometry method of choice for analysis of major and trace elements in bulk specimens, useful PIXE measurements can be made. A detailed review of the main considerations for thick-target PEXE provides guidance for trace analysis with known and unknown matrices and bulk analysis when the constituents are unknown. Campbell and Cookson also discuss the increased importance of secondary fluorescence and geometrical accuracy for bulk measurements. 

XRF nowadays provides accurate concentration data at major and low trace levels for nearly all the elements in a wide variety of materials. Hardware and software advances enable on-line application of the fundamental approach in either classical or influence coefficient algorithms for the correction of absorption and enhancement effects. Vendors software packages, such as QuantAS (ARL), SSQ (Siemens), X40, IQ+ and SuperQ (Philips), are precalibrated analytical programs, allowing semiquantitative to quantitative analysis for elements in any type of (unknown) material measured on a specific X-ray spectrometer without standards or specific calibrations. The basis is the fundamental parameter method for calculation of correction coefficients for matrix elements (inter-element influences) from fundamental physical values such as absorption and secondary fluorescence. UniQuant (ODS) calibrates instrumental sensitivity factors (k values) for 79 elements with a set of standards of the pure element. In this approach to inter-element effects, it is not necessary to determine a calibration curve for each element in a matrix. Calibration of k values with pure standards may still lead to systematic errors for unknown polymer samples. UniQuant provides semiquantitative XRF analysis [242]. 

Matrix absorption, secondary fluorescence and scattering phenomena limit sensitivity and precision in many cases, especially with dense matrices. The sensitivity falls off with atomic number elements with Z < 15 are particularly difficult to analyse. Analysis is characteristic of surface layers (5-500 pm depth) only for a solid specimen. Instruments are often large, complicated and costly. 

In addition to absorption problems, measurements will be affected by secondary fluorescence and scattered radiations which will enter the detector and increase the general background. Detection limits under optimum conditions (a heavy element in a light matrix) may be as low as 10 ppm. Quantitative analysis is however difficult below the 20-100 ppm region if a reasonable precision (5% or better) is to be obtained. 

Fluorescence occurs when radiant energy is absorbed and then, almost instantly, some of the energy is re-emitted, usually at a longer wavelength. Primary fluorescence (autofluorescence) occurs in flavo-proteins (13), plant cell wall materials such as lignin (7), and in flagella (14). Secondary fluorescence is when a material binds a fluorescent dye... 

Quantitation is usually achieved by comparing the X-ray yields from the sample with yields obtained from standards. The ease with which measurements can be interpreted quantitatively depends on the sample. As illustrated in Fig. 7.7, the volume that is activated by the 10-100 keV electron beam has the shape of a pear with typical dimensions of a few pm. As a consequence, X-rays formed in the interior may be absorbed on their way out, and may stimulate the emission of photoelectrons, Auger electrons and again X-rays. The latter process, secondary fluorescence, can lead to an overestimate of the concentrations. For example, if the specimen is a bulk Fe-Ni alloy, Ni Ka radiation is adsorbed by iron and causes... 

In bulk samples, X-ray yields need to be adjusted by the so-called "ZAF" correction. Z stands for the element number (heavier elements reduce the electron beam intensity more than lighter elements, because they are more efficient back-scatterers), A for absorption (different elements have different cross sections for X-ray absorption), and F for secondary fluorescence (the effect described above). Corrections are much less important when the sample is a film with a thickness of 1 pm or less, because secondary effects are largely reduced. The detection limit is set by the accuracy with which a signal can be distinguished from the bremsstrahlung background. In practice, this corresponds to about 100 ppm for elements heavier than Mg. 

XRF spectrometry is based on the principle that primary X-rays (from an X-ray tube or radioactive source) are incident upon a sample and create inner shell (K, L, M) vacancies in the atoms of the surface layers. These vacancies de-excite by the production of a secondary (fluorescent) X-ray whose energy is characteristic of the elements present in the sample. Some of these characteristic X-rays escape from the sample and are counted and their energies measured. Comparison of these energies with known values for each element (e.g., Van Grieken and Markowicz 1993, Parsons 1997) allow the elements present in the sample to be identified and quantified. 

In principle, the difference between X-ray fluorescence spectrometry and electron-probe microanalysis lies in the fact that the analytical information is provided, in the first case, by secondary, fluorescence X-rays, and in the second by primary X-rays, emitted as a result of the impact of the electron beam on the sample s electrons. 

Add the appropriate amount of secondary fluorescent antibody diluted in PBSG with 10% bovine serum albumin (see Note 2). 

Figure 4. Primary and secondary vitrinite fluorescence (absolute units) (a) primary fluorescence in U-V irradiation, (b) secondary fluorescence in blue-light irradiation.

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