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Isotope dilution assay in peptide quantification: The challenge of microheterogeneity of internal standard
Author(s) -
Stoyanov Alexander V.,
Rogatsky Eduard,
Stein Daniel,
Connolly Shawn,
Rohlfing Curt L.,
Little Randie R.
Publication year - 2013
Publication title -
proteomics – clinical applications
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.948
H-Index - 54
eISSN - 1862-8354
pISSN - 1862-8346
DOI - 10.1002/prca.201200130
Subject(s) - isotope dilution , analyte , chromatography , chemistry , peptide , dilution , amino acid analysis , isotope , quantitative analysis (chemistry) , amino acid , mass spectrometry , biochemistry , physics , thermodynamics , quantum mechanics
Isotope dilution analysis allows quantitation of elements and different compounds in complex mixtures. The quantitation is based on a known amount of reference material (internal standard, IS ) added to a sample that makes the result critically dependent on the value assigned to the standard. In the case of peptides, IS concentration is determined by nitrogen and amino acid analysis while purity is normally assessed by methods such as chromatography or electrophoresis that might not be able to detect many possible amino acid modifications, either naturally occurring or chemically induced. Microheterogeneity of the IS , if it is not accounted for when assigning a reference value to the standard, results in highly overestimated values in target analyte quantitation. In this viewpoint article, we illustrate the problem of internal standard microheterogeneity by analyzing synthetic human C ‐peptide labeled analogs.