Targeted Proteomics Analysis

Introduction of PRM

PRM (Parallel Reaction Monitoring) technology is a targeted verification proteomics technology based on mass spectrometry. Through selective detection of specific peptides or target peptides, targeted relative or absolute quantification of target proteins or modified peptides can be achieved . Its advantage lies in overcoming the difficulty of antibody preparation and the difficulty of buying antibodies, and it perfectly replaces antibody-based Western blot, ELISA and other technologies. PRM can be used for the verification of wide-screen data, such as genome, transcriptome, quantitative proteome, target protein or peptide obtained by modifying the proteome, to achieve relative or absolute quantitative analysis of protein or peptide.

Fig 1.Workflow for highly multiplexed, RT scheduled sPRM assay development. (Birgit S, et al.2015)

Technical Principle

PRM technology is based on a high-resolution, high-precision mass spectrometer (such as Q-Exactive HF-X). First, it uses the selective detection capability of a quadrupole mass analyzer to selectively detect the precursor ion information of the target peptide. Then it fragments in the HCD collision cell, and finally enters the high-resolution, high-quality precision Orbitrap analyzer to detect all fragments in the selected precursor ion window. In this way, the target protein or peptide in the complex sample can be accurately analyzed.

Reference

1. Birgit S, et al. Multiplexed, scheduled, high-resolution parallel reaction monitoring on a full scan QqTOF Instrument with Integrated data-dependent and targeted mass spectrometric workflows[J]. Analytical chemistry, 2015, 87( 20):10222.

Introduction of MRM

Multiple reaction monitoring (MRM) technology was also called selective reaction monitoring (SRM) in the early days. This technology is essentially a scanning mode of mass spectrometry. Based on the specific parent ion and product ion pairs of the target protein, signals that conform to the target ion rules are selected to remove signal interference that does not conform to the rules. Finally, a high-sensitivity, high-accuracy and specific targeted protein quantitative analysis is performed.

Advantages of MRM

High sensitivity. After two-stage mass spectrometry, the ions that are consistent with the target ion are selected to eliminate interfering ions, which greatly improves the signal-to-noise ratio and improves the accuracy of detecting target ions.

With high throughput, more than 200 proteins can be identified at one time.

Absolute quantitative analysis can be performed without relying on antibodies.

Identify low-abundance proteins with high precision, with a quantitative range spanning 4 orders of magnitude.

Technical Principle

MRM mass spectrometry analysis mainly includes three stages.

1. First-level mass spectrometry scans to screen out parent ions with the same specificity as the target molecule;

2. Collision to fragment precursor ions and remove interfering ions;

3. Collect only the mass spectrum signal from the selected specific ion.