Isobaric Dimethyl Labeling for Multiplex Proteome Quantification
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current multiplex proteome quantification methods face challenges with low signal-to-noise ratio, sensitivity, data analysis complexity, and limited throughput, particularly in MS1-based methods, while MS2-based methods improve accuracy and dynamic range but still fall short in throughput.
Innovation Solution
A multiplex proteome quantification method using isobaric dimethyl labeling, where peptides are labeled at the N-terminal and C-terminal under different pH conditions, utilizing specific reagents to achieve sextuple labeling and quantification by fragment ions in MS2, allowing for high labeling efficiency and selectivity, and simultaneous analysis of up to six samples.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If MS1-based quantitative method is used, then throughput is improved, but signal-to-noise ratio and sensitivity deteriorate
Solution Approach 1:
The patent segments the quantification process into two distinct stages: MS1 scan for high-throughput sample identification and MS2 scan for high-precision quantification. This segmentation allows the system to leverage the speed of MS1 while achieving the accuracy of MS2, resolving the contradiction between throughput and signal-to-noise ratio.
Solution Approach 2:
The patent introduces an intermediary approach by using MS1 data as a preliminary filter that guides subsequent MS2 analysis. The MS1 scan acts as an intermediary step that identifies candidate peptides, which then undergo detailed quantification in MS2, thereby improving overall throughput without sacrificing measurement precision.
2Measurement precision
If MS2-based quantitative method is used, then signal-to-noise ratio and quantitative accuracy are improved, but data analysis complexity increases
Solution Approach 1:
The patent performs preliminary action by conducting MS1 scans before MS2 scans. The MS1 data is processed and used to guide the selection of peptides for MS2 analysis, which simplifies the subsequent data analysis by pre-filtering and organizing the data in a structured manner.
Solution Approach 2:
The patent adds a temporal dimension to the data analysis by processing MS1 and MS2 data in sequence rather than simultaneously. This dimensional approach allows complex MS2 data to be analyzed in manageable stages, reducing overall computational complexity while maintaining high quantitative accuracy.
3Quantity of substance
If multiplex labeling is increased, then quantitative coverage is improved, but MS1 spectrum complexity increases
Solution Approach 1:
The patent extracts the quantification information from the complex MS1 spectrum and relocates it to the MS2 spectrum. By taking out the quantitative measurements from the crowded MS1 space and performing them in the more resolved MS2 space, the method maintains high quantitative coverage while avoiding MS1 spectrum complexity.
Solution Approach 2:
The patent moves the quantification measurement from the mass-to-charge ratio dimension in MS1 to the fragment ion dimension in MS2. This dimensional transition allows multiple labeled samples to be quantified simultaneously without increasing MS1 spectrum complexity, as the quantification occurs in the additional spectral dimension provided by fragmentation.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This method achieves high labeling efficiency and selectivity, wide quantitative dynamic range, and high throughput, with improved accuracy and precision, effectively reducing MS2 spectrum complexity and enabling efficient simultaneous analysis of multiple samples.
Implementation Method 1
dimethyl labeling is carried out on the peptide N-terminal and C-terminal of Lys-C digests
Implementation Method 2
LC-MS analysis. The mass spectrometry comprises Orbitrap, TOF and FT-ICR
Implementation Method 3
the intensity values of the fragment ions of a, b and y which are simultaneously shown in the sextuple labeling of each labeled peptide on the MS2 are extracted
Data Source
AI summary
A multiplex proteome quantification method based on isobaric dimethyl labeling implements dimethyl labeling of peptide N-terminal in an acidic condition and C-terminal in an alkaline condition one after another by means of Hall the property that a dimethylation reaction has different rates on an amino group at the peptide N-terminal and an amino group on a Lysine side chain at the peptide C-terminal in the acidic condition. Multiplex labeling of peptide samples is implemented by means of the organic combination of various isotope forms of a dimethyl labeling reagents. The mass-to-charge ratios in MS1 of peptides after multiplex labeling are completely the same, the mass-to-charge ratios of the fragment ions in MS2 are different, and multiplex quantitative analyses are carried out by extracting the intensity values of corresponding fragment ions in the MS2.