Multiple Reaction Monitoring

Multiple Reaction Monitoring (MRM), also known as Selected Reaction Monitoring (SRM), is a targeted tandem mass spectrometry (MS/MS) scan mode on triple quadrupole instruments. In lab reports or Certificates of Analysis (COAs), MRM is used to confirm the identity of a specific known substance and quantify its concentration. It is widely used for measuring peptides, proteins, drugs, hormones, and environmental toxins, often with liquid chromatography (LC-MS/MS).

How MRM uses dual mass filters for high specificity

MRM achieves high specificity by applying two independent mass filters in sequence. This makes the signal more precise than single-stage mass spectrometry because it helps distinguish the target substance from background noise or similar-looking molecules. The specific precursor-to-fragment pair is called an MRM transition. Because MRM is targeted, the laboratory must already know the molecule's mass and fragmentation behavior before starting the test.

The MRM workflow has three stages:

1. Q1 (first mass filter) selects the target molecule by its mass.
2. Q2 (collision cell) fragments the molecule into smaller pieces.
3. Q3 (second mass filter) selects a specific fragment for detection.

Limitations of MRM in general purity testing

MRM is not a general purity screen and will not detect substances it was not specifically programmed to find. It does not identify unprogrammed impurities. Consequently, a percentage purity figure typically cannot be determined by MRM alone and requires additional orthogonal methods. For discovering unknown impurities, untargeted LC-HRMS is the more appropriate analytical approach.

Sources

1. Multiple Reaction Monitoring-based, Multiplexed, Absolute Quantitation of 45 Proteins in Human Plasma
2. MRM-based LC-MS method for accurate C-peptide quantitation
3. Methods for Peptide and Protein Quantitation by Liquid Chromatography-Multiple Reaction Monitoring Mass Spectrometry - PMC
4. An Introduction to Advanced Targeted Acquisition Methods
5. Automated Detection of Inaccurate and Imprecise Transitions in Peptide Quantification by Multiple Reaction Monitoring Mass Spectrometry - PMC
6. [PDF] M10 BIOANALYTICAL METHOD VALIDATION AND STUDY ... - FDA
7. Quantitative analysis of peptides and proteins in biomedicine by targeted mass spectrometry
8. Identification and accurate quantification of structurally related peptide impurities in synthetic human C-peptide by liquid chromatography-high resolution mass spectrometry - PubMed
9. Bioanalytical Method Validation - Guidance for Industry
10. Current Role and Potential of Triple Quadrupole Mass Spectrometry in Biomedical Research and Clinical Applications - PMC
11. Targeted Quantitation of Proteins by Mass Spectrometry - PMC
12. Parallel Reaction Monitoring: A Targeted Experiment Performed Using High Resolution and High Mass Accuracy Mass Spectrometry
13. Multiple and Selective Reaction Monitoring Using Triple ...
14. Liquid Chromatography-High Resolution Mass Spectrometry for Peptide Drug Quality Control - PMC
15. Advancing the sensitivity of selected reaction monitoring-based targeted quantitative proteomics
16. Basic design of MRM assays for peptide quantification - PubMed
17. LC-MS/MS for protein and peptide quantification in clinical chemistry - PubMed
18. Large-Scale Interlaboratory Study to Develop, Analytically Validate and Apply Highly Multiplexed, Quantitative Peptide Assays to Measure Cancer-Relevant Proteins in Plasma