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Absolute quantitation (AQUA) of peptides

In short: peptides or peptide fragments of the protein that needs to be quantified are mixed with their stable isotope-labeled (13C15N) counterparts of a known amount. The intensity ratios between these two peptides are measured and the absolute amount of the non-labeled peptide is calculated.  The Proteomics Resource Center provides the synthesis of heavy stable isotope-labeled peptides. Please note that utilizing aliphatic amino acids (Leucine, Alanine, and Valine) for introduction of the heavy stable isotope atoms is far less expensive than utilizing amino acids which require side chain reactive group protection (Arginine and Lysine).

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Include example figures.

 

How absolute is absolute – not very. CoCONFRAC.

Depending on digestion….