Toward Système International d'Unité-traceable protein quantification: from amino acids to proteins

Anal Biochem. 2008 May 15;376(2):242-51. doi: 10.1016/j.ab.2008.02.010. Epub 2008 Feb 19.

Abstract

Here we present a demonstration of the proof of principle that absolute concentration of a protein within a mixture of other proteins can be measured with SI traceability. The method used was based on tryptic digestion of a protein followed by quantification using double exact matching isotope dilution mass spectrometry (IDMS) of the peptides released. To provide full SI traceability to measurements of protein concentration we demonstrated a method of SI traceable peptide quantification in which the peptide standards used were quantified by an amino acid analysis method that incorporated double exact matching IDMS and amino acid standards of known purity. The concentration of the protein was therefore determined based upon the concentration of tryptic peptides, which in turn had been quantified based upon amino acid standards. This allowed fully SI-traceable measurements of protein concentration to be made. Important caveats in the implementation of this approach are also discussed and examples of how these can have detrimental effects on the measurements are shown.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acids / analysis*
  • Amino Acids / chemistry
  • Chromatography, Liquid
  • International System of Units*
  • Isotope Labeling / methods
  • Mass Spectrometry
  • Proteins / analysis*
  • Proteins / chemistry
  • Proteomics / methods

Substances

  • Amino Acids
  • Proteins