Method for monitoring and qualification of 15N metabolic labeled protein in combination with mass spectrum parallel reaction

Abstract

The invention discloses a method for monitoring and qualification of 15N metabolic labeled protein in combination with mass spectrum parallel reaction. 15N metabolic labeled internal reference proteincan accurately reflect the structure and the state of endogenous protein to be detected, has the same enzymolysis efficiency with the protein to be detected, solves the problem that internal reference peptide ion strength signals are affected by protease digestion, co-elute matrix, sample composition and instrument status during mass spectrum selection, tracking and quantification to cause nonreproducibility, and meanwhile improves the efficiency of monitoring and qualification of mass spectrum parallel reaction; in addition, the method can produce various quantitative polypeptide used for parallel reaction monitoring, improves the qualification accuracy, reduces the cost for synthesizing polypeptide, can be applied to quantitative proteomic research and detection of protein markers, andhas great help in medical diagnosis, environmental health, food safety inspection and judicial expertise.

Description

technical field [0001] The invention relates to the analytical field of mass spectrometry detection, especially a 15 A method for the quantification of N metabolically labeled proteins combined with mass spectrometry for parallel reaction monitoring. Background technique [0002] Mass spectrometry parallel reaction monitoring (Parallel reaction monitoring, PRM) technology, as a mass spectrometry detection method, has the advantages of strong specificity, high sensitivity, high accuracy, good reproducibility, wide linear dynamic range, automation and high throughput. Advantages, these qualities can meet the urgent needs of many research fields today. Real-time quantitative monitoring through PRM technology can be used for pharmacokinetics, clinical diagnosis, illicit drug inspection, food and cosmetics and other industrial quality control, environmental testing, agricultural botany research, and the discovery of metabolomics and proteomics biomarkers, etc. Related research....

AI Technical Summary

Problems solved by technology

First, this method requires preliminary experiments and extensive data analysis to determine the target peptide for synthesis

Second, the protease digestion process of different batches of samples can produce significant differences

The results showed that the synthetic peptides could not accurately reflect the absolute amount of endogenous protein measured, and the difference was often more than 10 times

Third, quantification is limited to endogenous peptides identical to synthetic peptides, rather than the entire protein sequence. Another disadvantage is that synthetic peptides are expensive compared to protein targets.

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