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Benefits of Stable Isotope Labelled Proteins
as Internal Standards in Quantitative LC-MS workflows

A stable isotope labelled (SIL) form of an analyte protein is widely regarded as the optimal internal standard (IS) for absolute quantification of proteins using LC-MS; SIL proteins are often cited as being the “gold standard IS”.

The benefit of using a SIL protein as an internal standard is that shares the same physiochemical behaviour as the target protein analyte. This means that the IS can be introduced at the start of the sample preparation workflow and will be processed along with the analyte protein throughout the entire analytical procedure (see schematic above). The addition of SIL-proteins at the start of the sample extraction can control for variations due to immunoaffinity or other isolation techniques, enzymatic digestion, other pre-analytical treatments as well as the LC and mass spectrometric ionization.

EMA guidelines recommend the use of a SIL-protein IS whenever possible for LC-MS/MS methods. Beyond this, there is a growth in scientific literature that strongly supports the use of full length, recombinant SIL proteins as internal standards and selected publications discussing the benefits of using SIL proteins in quantitative LC-MS workflows are detailed above. Significantly, these citations illustrate that the addition of a SIL protein as an IS at the start of the sample preparation increases accuracy and precision of the quantitative LC-MS assay when compared to other IS approaches e.g. SIL peptides or SIL extended peptides.

Our stable isotope labelled (and unlabelled)  full length
recombinant protein products are especially suitable for:

  • Biomarkers evaluation and validation > Labelled proteins provide the most reliable, reproducible and accurate quantification data for quantitative LC-MS work
    > Products are ready-to-use and packaged to be highly stable.
    > To use requires a simple modification to the workflow by spiking a controlled concentration of the SIL protein minimising sample processing variation and reduces assay development time
  • Development of Certified Reference Materials
  • Pharmacokinetics studies of biotherapeutics > Promise Proteomics has an extensive experience in producing labelled therapeutic proteins and antibodies for use in PK/PD studies using LC-MS.
    > Promise has participated in collaborative R&D projects with pharmaceutical companies based on our high-level expertise in monoclonal antibody quantitation.
  • Quantification of therapeutic monoclonal antibodies in clinical research studies

Key Benefits of Promise Proteomics
stable isotope labelled proteins

  • High protein purity & isotopic incorporation
  • Flexibility in peptide selection
  • Same sequence as native protein
  • Equivalent digestion kinetics to target
  • Identical to analyte in sample processing
  • For use in complex sample matrices
  • Highly stable
  • Catalogue & custom options available

Publications Citing the Benefits of using Full Length Stable Isotope
Labelled Recombinant Proteins as Internal Standards in Quantitative LC-MS