Ratiometric comparison

When a standard is used for quantification of the concentration of an analyte, many researchers simply use a formula of ratiometric comparison ... 

This approach has been used to quantify individual species of over 40 lipid classes directly from lipid extracts of biological samples with or without derivatization [16, 18, 29, 37]. For example, in the analysis of SM species present in lipid extracts of mouse cortices, which were treated with lithium methoxide as previously described [38], the quantities of lithiated N18 l SM at miz 735.5, N18 0 SM at m/z 737.5, and N24 l SM at m/z 819.6 were accurately measured with ratiometric comparisons to the selected internal standard (i.e., N14 0 SM at m/z 653.5) after de-isotoping of the mass spectrum acquired in the full MS mode (Trace a of Figure 14.2). The contents of all other low-abundant SM species were determined with the second step of quantitation utilizing the mass spectrum of NLS 183 (Trace b of Figure 14.2) or NLS213 (as previously demonstrated [20]) with N14 0, N18 l, and N24 l SM as internal standards. 

Quantification by ratiometric comparison with an internal standard is based on the ratio of the sum of the isotopologue intensities of a species to that of the internal standard. The fact is that the monoisotopic peak is the most intense peak in the isotopologue cluster of a lipid species for almost all lipids, and its intensity can therefore be determined more accurately compared to the intensities of other isotopic peaks of the species. Therefore, correcting for differential isotopologue distribution is based on the deduction of the intensity of each isotopologue of a species from the determined monoisotopic peak intensity. 

Vitha MF, Clarke RJ (2007) Comparison of excitation and emission ratiometric fluorescence methods for quantifying the membrane dipole potential. Biochim Biophys Acta-Biomembr 1768(1) 107—114... 

A detailed stndy of the combination of flnorescein boronic acid with diol-appended quenchers a-c and comparison with the fluorescence outputs of nonboron or nondiol-containing systems (i.e., fluorescein or methyl red were employed directly) revealed that the boronate ester formation results in enhanced quenching in each case, and that compound c is the best overall quencher. Nncle-osides were also shown to bind to the same fluorescein boronic acid derivative. While the quenching ability of each nucleoside tested was different, the same ratiometric quenching enhancement was observed in each case, sng-gesting similar binding affinities. 

In most examples presented above, the sensing function is based on changes in the intensity of one transition, which can be heavily affected by the quantity of the luminophore, excitation power, and the drifts of the optoelectronic system. Thus, the comparison of the emission of different samples based on the detected intensity may lead to erroneous conclusions. Although the measurements of quantum yields and/or lifetime are affected neither by the intensity of the excitation source nor by the probe concentration, they require a relatively long time and the computational treatment. The utilizing of the ratio between the intensity of two transitions of the same luminescent material, instead of only one transition, can overcome the main drawbacks of the intensity-based measurements of only one transition. The ratiometric... 

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