What is TMT?
TMT (Tandem Mass Tag) is an isotopic labeling technique used for quantitative analysis in proteomics. It employs isotopic tags for relative or absolute quantification of proteomes, allowing simultaneous comparison and analysis of multiple samples. The core of TMT technology is a series of isotopic labeling reagents that can covalently bind with proteins or peptides, forming isotopically labeled products detectable on a mass spectrometer.
1. The TMT tag consists of three components:
1. An active group that can bind to the terminal amino acids (usually lysine) of peptides;
2. A mass normalization region that can dissociate in the mass spectrometer;
3. A unique mass reporter region containing different isotopic compositions.
2. The basic steps of a TMT experiment are as follows:
1. Protein extraction:
Extract proteins from different samples.
2. Protein digestion:
Digest the extracted proteins, typically using trypsin, to generate peptides.
3. Peptide labeling:
React the peptides from each sample with different TMT tags to form labeled peptides.
4. Sample mixing:
Mix the labeled peptides together to form a combined sample.
5. Peptide separation:
Separate the peptides in the mixed sample using liquid chromatography (such as high-performance liquid chromatography, HPLC).
6. Mass spectrometry analysis:
Conduct mass spectrometry analysis (typically tandem mass spectrometry, MS/MS) on the separated peptides, detecting the reporter ions of the TMT tags in the mass spectra of fragment ions.
7. Data processing:
Calculate the relative or absolute abundance of proteins in different samples by comparing the intensity of reporter ions from different TMT tags.
3. Advantages of TMT technology:
1. High sensitivity:
Can detect low-abundance proteins;
2. Strong separation capability:
Can separate acidic/basic proteins, proteins smaller than 10KD or larger than 200KD, insoluble proteins, etc.;Broad application range:Can identify any type of protein, including membrane proteins, nuclear proteins, and extracellular proteins;
3. High throughput:
Can analyze 10 samples simultaneously, especially suitable for differential proteomics analysis of samples subjected to various treatment methods or durations.
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Related services:
Label-free quantitative proteomics analysis
TMT/iTRAQ/MultiNotch quantitative proteomics analysis
SWATH quantitative proteomics service
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