Isotope method validation

Jemal, M., Schuster, A., and Whigan, D. B. (2003). Liquid chromatography/tandem mass spectrometry methods for quantitation of mevalonic acid in human plasma and urine method validation, demonstration of using a surrogate analyte, and demonstration of unacceptable matrix effect in spite of use of a stable isotope analog internal standard. Rapid Commun. Mass Spectrom. 17, 1723-1734. 

It is important to first note that many classical clinical assays have traditionally measured one metabolite to detect one disease. Consequently, many of the rules of method validation were designed around this premise. MS/MS, as originally designed, detected two classes of compounds, amino acids and acylcamitines, in four to five different MS methods (known as scan modes such as neutral loss, precursor ion, or selected reaction monitoring), for approximately 500 distinct masses, more than 70 known compounds, and 20-30 stable isotope internal standards. How then did one approach such a complicated validation to gain acceptance as a reliable, useful method The answer is quite simple - start simply and compare to what was already established. 

In light of the obvious flaws in their evaluation, we once again contend that their experiments did not constitute a valid evaluation of om" wash kinetic procedures. Nor do we agree with them that their described isotope procedure is suitable for contamination/decontamination studies. As a matter of fact, it is likely that an inherent difficulty in their isotope procedures, which we shall describe shortly, may have created the observed difference between their first and second studies. It is also likely that insufficient attention was given to critical elements in their contamination/decontamination experiments. Since we have not described the latter in our previous publications, we will briefly simunarize these now and then comment on a general problem with their isotope method. 

Direct measurement of dietary zinc availability in humans requires development of the stable isotope tracer methodology. Several aspects of this integrated methodology are considered and briefly discussed. These are analytical isotopic measurement methodology, consequences of the finite precision of isotopic measurements, validation of in vivo measurements, and several aspects of biological labeling of human foods. It is shown that Radiochemical Neutron Activation Analysis provides a suitable method for accurate measurement of the stable isotopes Zn,... 

To evaluate the possible effects of isotopic fractionation on the validity or usefulness of the lead isotope method, a fractionation experiment was devised to simulate the probable ancient production methods. 

The kinetic isotope method was also applied to study whether the stepwise or direct route is valid for the dehydrogenation of the cyclohexane ring.[bi4,67] mentioned before, plotting specific radioactivities as a function of the contact time confirmed the predominance of direct pathway on Re and the stepwise dehydrogenation on chromia. 

Based on the data presented here and in the subsequent volumes of this report, the methodology employed in the generation of spent fuel isotopics and the criticality calculations using those isotopes is valid for bumup credit analyses. Further validation of the SCALE S AS2H sequence for generating spent fuel isotopics has been performed by comparison of calculated results with measured spent fuel chemical assay data. The criticality methods of CSAS/KENO V.a are validated against LWR-type fresh fuel critical experiments (both UO2 and MOX) in ref 19. 

In all these applications, isotope ratio data are produced, which are interpreted on an absolute or relative basis and which have an impact on our daily life, whether this is in science (e.g., age of an artifact), in society (e.g., provenance of food), or in public safety (e.g., neutron shielding in nuclear power plants). To ensure that these data are reliable and accurate, some specific requirements have to be fulfilled. The main requirement is that all these measurement results are comparable, which means that the corresponding results can be compared and differences between the measurement results can be used to draw further conclusions. This is only possible if the measurement results are traceable to the same reference [25]. This in turn can only be realized by applying isotopic reference materials (IRMs) for correction for bias and for validation of the analytical procedure. Whereas in earlier days only experts in mass spectrometry were able to deliver reproducible isotope ratio data, nowadays many laboratories, some of which may even have never been involved with mass spectrometry before, produce isotope ratio data using inductively coupled plasma mass spectrometry (ICP-MS). Especially for such users, IRMs are indispensable to permit proper method validation and reliable results. The rapid development and the broad availability of ICP-MS instrumentation have also led to an expansion of the research area and new elements are under investigation for their isotopic variations. In this context, all users require IRMs to correct for instrumental mass discrimination or at least to allow isotope ratio data to be related to a commonly accepted basis. 

Lee, W., McCabe, G.P., Martin, B.R., and Weaver, C.M. (2011) Validation of a simple isotope method for estimating true calcium ifactioiial absorption in adolescents. Osteoporos. Int., 22, 159-166. 

Isotopic methods take advantage of isotopes for labelling purposes. i.e. for tagging either atoms or molecules which thus can be detected and traced. The validity of such methods depends on the basic assumption that the various isotopic forms of any given element differ only as to their physical properties, while possessing the same chemical behaviour. It is similarly assumed that a molecule labelled with one or more isotopes of its elemental constitumits will behave exactly like the other unlabelled molecules of the same compound. For practical purposes, these assumptions hold true in the majority of cases and can be con-... 

The Advanced Seminars have been crucial for evaluating the utility and validity of existing methods and results, discussing potential improvements in methods, and setting priorities for future research. A brief summary of the advances made in previous seminars and future directions of research on chemical and isotopic techniques of diet reconstruction follows. 

There are a few developments on the horizon that will increase our ability to date bones and teeth reliability. Both y- and a-spectrometric methods can measure Pa/ U and °Th/U and concordance between dates calculated using the two can provide a measure of reliability. However, the discordance between the two is not very sensitive to different uptake regimes, and it is difficult to resolve, for example, bones that have undergone EU from those that have undergone LU with the analytical errors commonly encountered in measurements by y- and a-spectrometry. On the other hand, it has been shown recently that TIMS can measure both isotopic ratios with a precision usually better than 1% (Edwards et al. 1997). TIMS measurements of Pa/ U and °Th/U have yet to be routinely applied to dating fossil remains, but in the future, concordance between the two decay series will provide further evidence of the validity of a particular uptake model to a particular sample. 

A simple NMR technique, and arguably the most widely used and effective for hit validation, is the chemical shift perturbation method. In this approach, a reference spectrum of isotopically labeled target is recorded in absence and presence of a given test ligand (or a mixture of test ligands). Commonly, differences in chemical shift between free and bound protein target are observed in 2D [15N, 1H and/or 2D [13C, H] correlation spectra of a protein (or nucleic acid) upon titration of a ligand... 

The method development process with the multisorbent plate consists of three steps. In step 1, the sorbent chemistry and the pH for loading, washing, and elution are optimized. In step 2, optimization of the percentage organic for wash and elution and the pH of the buffer needed is carried out. Step 3 is validation the method developed from the results of the previous two steps is tested for linearity, limits of detection, quantitation of recovery, and matrix effects using a stable isotope-labeled analyte as an IS. 

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