Binding absolute accuracy

The absolute accuracy of binding energies between different instruments and laboratories varies even for noble metals significantly more than the numerical accuracy with which the peak position in a spectrum can be determined (more than the usually quoted 0.1 eV). This is independent of the method of spectrometer calibration and sample preparation. 

For an extraction of precise energy values from measured absorption edges additional information is required one may, for example, take the energy differences between the distinct bound and continuum levels from optical spectroscopy data of the next element (optical match, Z + 1 rule). With this method inner-shell binding energies have been determined with absolute accuracies of about The experimental binding energies... 

Finally, changes in ion binding and other cell parameters in disease states can also complicate the interpretation of the data and result in errors. Again, while it is possible to make many microelectrode measurements precisely and then to use deviations from normal values as a means of clinical diagnosis, little is really known at the present time in terms of the absolute accuracy of these measurements. 

Values of 20% (25% at lower limit of quantification [LLOQ]) are recommended as default acceptance criteria for accuracy and inter-batch precision for ligand binding in practical use. Precision and accuracy should be established by analyzing four sets of QC samples at LLOQ, low, medium, and high levels in duplicate in six different batches during method development. In addition, a second proposed criterion for method acceptance takes into account the sum of inter-batch precision and the absolute value of accuracy be <30%. During practical application of such assays, the 4-6-30-rule may be applied - that is, for each batch four out of six QC samples must be within 30% of nominal concentration, but the two failed QC samples may not be at the same level. 

For each in-study run, the standard curve must satisfy criteria described in the standard-curve section however, run acceptance is based primarily on the performance of the QC samples. When using total error for ligand binding assays of macromolecules, the run acceptance criteria recommended in the precision and accuracy section requires that at least four of six (67%) QC results must be within 30% of their nominal values, with at least 50% of the values for each QC level satisfying the 30% limit. The recommended 4-6-30 rule imposes limits simultaneously on the allowable random error (imprecision) and systematic error (mean bias). If the application of an assay requires a QC target acceptance limit different than the 30% deviation from the nominal value, then prestudy acceptance criteria for precision and accuracy should be adjusted so that the limit for the sum of the interbatch imprecision and absolute mean RE is equal to the revised QC acceptance limit. 

Pro The Lowry assay exhibits the best accuracy with regard to absolute protein concentrations due to the chemical reaction with polypeptides. This contrasts with the other two methods, which, as dye binding assays, exhibit more variation depending on the different reactivity of the given proteins (standards as well as samples). 

In most cases, however, accuracy is critical. In measuring the stoichiometries of ligand binding to proteins it is essential to know the true molecular weight and concentration of the protein. We shall therefore now discuss a number of practical aspects that should allow the absolute calibration of assays for macromolecules, particularly proteins and DNA... 

A possible explanation for the absolute shift of the 3d core hole binding energies may be found in the LDA itself. Although the Hedin-Lundqvist (1971) form of the exchange and correlation potential produces good results for the valence electrons, it is questionable whether it describes a 3d core state with the same accuracy. In... 

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