What Advanced Techniques are Used for Absolute Quantification in Proteomics?

One important direction of proteomics research is the quantitative analysis of proteins. Traditional relative quantification methods provide information on the relative abundance of proteins, but cannot accurately measure the absolute abundance of proteins. To overcome this limitation, researchers have developed many advanced techniques for absolute quantification of proteomics. This article will detail the advanced techniques commonly used in proteomics research for absolute quantification of proteomics, revealing the latest developments and applications of proteomic quantification for readers.

 

Isotope Labeling Quantitation

1. Stable Isotope Labeling with Amino Acids in Cell Culture (SILAC)

This technique quantifies proteins by using isotope-labeled amino acids in cell culture.

 

2. Isobaric Tags for Relative and Absolute Quantitation (iTRAQ)

This technique uses isotope-labeled compounds to label protein samples, then uses mass spectrometry to achieve absolute quantification of proteins.

 

Parallel Reaction Monitoring Quantitation

1. Selected Reaction Monitoring (SRM)

This technique uses mass spectrometry to selectively monitor specific peptide segments, achieving absolute quantification of proteins.

 

2. Parallel Reaction Monitoring (PRM)

Similar to SRM, this technique uses mass spectrometry to simultaneously monitor multiple specific peptide segments, achieving absolute quantification of proteins.

 

Protein Abundance Calculation

1. Top3 Calculation

This technique calculates the relative and absolute abundance of proteins based on the peptide signal of the top three peaks in mass spectrometry analysis.

 

2. Protein Abundance Index (PAI)

This technique estimates relative and absolute protein abundance by comparing the spectral count of a protein to its theoretical spectral count.

 

Parallel Reaction DNA Quantitation

Digital PCR (dPCR): This technique achieves absolute DNA quantification by dividing a DNA sample into many small reaction volumes.

 

Considerations for Selecting an Appropriate Technique

1. Sample Properties

Different types of samples may require different quantification methods, such as cell culture supernatant, tissue samples, or bodily fluid samples.

 

2. Research Purpose

Depending on the purpose of the research, such as protein identification, quantification, or protein interaction studies, choose the appropriate quantification method.

 

3. Technical Conditions

Consider the laboratory equipment and technical level, and choose a suitable technique for absolute protein quantification.

 

Absolute quantification of proteomics is of great significance in biopharmaceutical research. With the continuous development of technology, advanced techniques such as isotope labeling quantitation, parallel reaction monitoring quantitation, protein abundance calculation, and parallel reaction DNA quantitation provide effective means for absolute quantification of proteomics. When choosing the right quantification technique, it is necessary to consider factors such as sample properties, research purpose, and experimental conditions. The application of these advanced techniques will provide more accurate and reliable quantification data for proteomics research, promoting the progress of biopharmaceutical research and development and clinical applications.