Related Experiment Video
Updated: Apr 1, 2026
Quantification of Proteins Using Peptide Immunoaffinity Enrichment Coupled with Mass Spectrometry
Published on: July 31, 2011
Multiplexed Quantification of Proglucagon-Derived Peptides by Immunoaffinity Enrichment and Tandem Mass Spectrometry
Anita Y H Lee1, Derek L Chappell1, Monika J Bak2
1Translational Biomarkers, Merck Research Laboratories, Rahway, NJ;
Background:
Proglucagon-derived peptides (PGDPs), which include glucagon-like peptide (GLP)-1, glucagon, and oxyntomodulin, are key regulators of glucose homeostasis and satiety. These peptide hormones are typically measured with immuno-based assays (e.g., ELISA, RIA), which often suffer from issues of selectivity.
Methods:
We developed a multiplexed assay for measuring PGDPs including GLP-1 (7-36) amide, GLP-1 (9-36) amide, glucagon, and oxyntomodulin by mass spectrometry and used this assay to examine the effect of a meal tolerance test on circulating concentrations of these hormones. Participants fasted overnight and were either given a meal (n = 8) or continued to fast (n = 4), with multiple blood collections over the course of 3 h. Plasma samples were analyzed by microflow immunoaffinity (IA)-LC-MS/MS with an isotope dilution strategy.
Results:
Assay performance characteristics were examined and established during analytical validation for all peptides. Intra- and interassay imprecision were found to be 2.2%-10.7% and 6.8%-22.5%, respectively. Spike recovery was >76%, and dilution linearity was established up to a 16-fold dilution. Immediately after the meal tolerance test, GLP-1 and oxyntomodulin concentrations increased and had an almost identical temporal relationship, and glucagon concentrations increased with a slight delay.
Conclusions:
IA-LC-MS/MS was used for the simultaneous and selective measurement of PGDPs. This work includes the first indication of the physiological concentrations and modulation of oxyntomodulin after a meal.
More Related Videos
Related Concept Videos
01:33Peptide Identification Using Tandem Mass Spectrometry
This technique helps gather information regarding the protein from which the peptide was obtained and to study the peptides’ amino acid sequence. Identifying peptides from a complex mixture is an important component of the growing field of...
01:24Glucagon-like Receptor Agonists
GLP-1, when administered in high doses intravenously, triggers insulin secretion, inhibits glucagon release, slows gastric emptying, reduces food intake, and restores normal insulin secretion. However, its rapid inactivation by...