Utilizing enzymatic digestion procedures in the bioanalytical laboratory
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Updated: Mar 29, 2026
Evaluation of Protein–Protein Interactions using an On-Membrane Digestion Technique
Published on: July 19, 2019
Matthew Szapacs1, Thomas Mencken1, Jon Williams2
1Bioanalytical Science & Toxicokinetics, Drug Metabolism & Pharmacokinetics, Platform Technology & Science, GlaxoSmithKline, 709 Swedeland Road, King of Prussia, PA 19406, USA.
Abstract:
In recent years, the use of LC-MS technologies in the bioanlytical laboratory for quantitation of peptide/protein biomarkers and biotherapeutics has increased dramatically. The increased interest is due to the improvement in sensitivity of MS instruments over the last 5-10 years, as well as its proven ability to overcome some common issues associated with immunoassay, namely selectivity and reagent availability. However, large proteins (>10 kDa) chromatograph and ionize poorly. To overcome this challenge, LC-MS/MS workflows for proteins larger than 10 kDa utilize enzymatic digestion procedures with subsequent quantitation of one or more of these enzymatically derived peptides to act as a surrogate for the intact protein. Here, recommendations of digestion technique and potential internal standards are summarized.
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