iTRAQ/TMT/MultiNotch Quantitative Proteomics Service

What Is iTRAQ/TMT/MultiNotch Quantitative Proteomics Service

iTRAQ, Isobaric Tags for Relative and Absolute Quantitation, and TMT, Tandem Mass Tags, are isobaric labeling techniques used for multiplexed quantitative proteomics. Both approaches use chemically encoded tags that generally contain three functional components: a reporter group, a balance group, and a reactive group.

 

The reactive group labels peptides, while the reporter and balance groups maintain an identical overall tag mass. As a result, peptides labeled with different channels appear at the same precursor mass in the MS spectrum. During MS/MS fragmentation, the reporter groups generate channel-specific ions whose intensities provide quantitative information on relative peptide abundance across samples.

 

MultiNotch extends isobaric quantitative proteomics with additional mass spectrometric strategies designed to improve quantitative accuracy. Together, these technologies form the basis of the iTRAQ/TMT/MultiNotch Quantitative Proteomics Service for comparative protein abundance analysis.

Chen, X. et al. Genomics Proteomics Bioinformatics. 2021.

Figure 1. iTRAQ/TMT Labeling Strategy for Clinical Proteomics

When This Service Is a Good Fit

The iTRAQ/TMT/MultiNotch Quantitative Proteomics Service is well suited to studies that require direct comparison of protein abundance across multiple predefined experimental groups. Depending on the labeling format, up to 10 samples can be incorporated into a multiplexed LC-MS/MS workflow.

 

Typical study designs include treatment-versus-control comparisons, multiple treatment conditions, and comparisons across developmental stages. Isobaric multiplexing is particularly useful when researchers want to analyze several related samples together and obtain relative proteomic changes within the same quantitative experiment.

 

For studies involving complex samples or demanding quantitative comparisons, MultiNotch-based strategies can provide an additional option when improved quantitative accuracy is important.

iTRAQ/TMT/MultiNotch Quantitative Proteomics Service at MtoZ Biolabs

MtoZ Biolabs provides an integrated iTRAQ/TMT/MultiNotch Quantitative Proteomics Service covering protein reduction and digestion, peptide labeling, LC-MS/MS analysis, mass spectrometry data processing, and downstream bioinformatics analysis.

 

The analytical platform includes Orbitrap Exploris 480 and Orbitrap Fusion Lumos mass spectrometers coupled with LC separation. For studies that require additional control of quantitative interference, MtoZ Biolabs can also incorporate MultiNotch MS strategies using MS3 or proton-transfer reaction approaches.

 

Researchers can provide their study objective and protein samples, and the MtoZ Biolabs team can complete the subsequent analytical workflow from sample preparation through quantitative proteomic data analysis and biological interpretation.

Analysis Workflow

1. Protein Extraction and Digestion

Proteins are extracted from samples, followed by reduction, alkylation, and enzymatic digestion into peptides.

 

2. iTRAQ or TMT Peptide Labeling

Peptides from different samples are labeled with distinct iTRAQ or TMT reporter channels.

 

3. Sample Pooling and Peptide Fractionation

Labeled peptides are combined and fractionated to reduce sample complexity.

 

4. LC-MS/MS Analysis

Peptides are separated by LC and analyzed by high-resolution mass spectrometry, with MultiNotch MS3 used when required.

 

5. Protein Identification and Relative Quantification

Peptides and proteins are identified, and reporter ion signals are used for relative protein quantification.

 

6. Bioinformatics Analysis and Biological Interpretation

Differential proteins are evaluated through statistical analysis, visualization, functional annotation, and enrichment analysis.

Griss, J. et al. J Proteome Res. 2019.

Figure 2. Scheme of the Entire Workflow for iTRAQ and TMT

Why Choose MtoZ Biolabs?

1. Multiplexed Comparison of Up to 10 Samples

iTRAQ and TMT workflows support efficient comparison of multiple predefined groups within one quantitative experiment.

2. MultiNotch MS3 for Interference-Prone Samples

MultiNotch MS3 can reduce reporter ion interference and improve quantitative accuracy in complex proteomic backgrounds.

3. Orbitrap Platforms Optimized for Isobaric Quantification

Orbitrap Exploris 480 and Orbitrap Fusion Lumos provide high-resolution MS performance for TMT-based proteomics.

4. One-Stop Service

MtoZ Biolabs provides integrated support from protein preparation and labeling to LC-MS/MS analysis, quantitative data processing, and bioinformatics.

5. Complex Sample Support

Established workflows support tissues, cells, microorganisms, body fluids, and other complex biological matrices.

Applications

The iTRAQ/TMT/MultiNotch Quantitative Proteomics Service can support a wide range of comparative proteomics studies, including:

1. Disease Mechanism Research

Identify differential proteins and pathways associated with disease-related biological changes.

2. Drug and Treatment Response

Compare proteomic changes across treated, untreated, dose, or time-point groups.

3. Developmental Biology

Characterize protein abundance changes across developmental stages or differentiation processes.

4. Biomarker Candidate Discovery

Screen differential proteins for further validation in biomarker-oriented research.

5. Functional Genomics

Evaluate proteomic changes associated with gene knockout, knockdown, overexpression, or mutation.

6. Systems Biology

Integrate quantitative protein changes with pathway and network analysis to support biological interpretation.

7. Plant and Microbial Research

Investigate proteomic responses to environmental conditions, stress, growth stages, or experimental treatments.

Sample Submission Suggestions

1. Sample Amount

The following quantities are general recommendations for the iTRAQ/TMT/MultiNotch Quantitative Proteomics Service. Exact requirements may vary with sample type and study design.

Sample Type	Recommended Quantity	Minimum Quantity
Animal Tissue	20–100 mg	10–50 mg
Plant Tissue	2–5 g	1–2 g
Cell Pellet	1 × 10⁷ cells	2 × 10⁶ cells
Microbial Samples	50 mg or 50 μL	20 mg or 20 μL
Serum/Plasma	10-50 μL	5-25 μL
Urine	1 mL	0.5 mL
Cerebrospinal Fluid	0.1 mL	0.05 mL
Protein Solution	20 μL	10 μL

2. Storage and Shipping

Samples should be stored at -80°C when applicable and shipped on dry ice. Avoid repeated freeze-thaw cycles and keep collection, processing, and storage conditions consistent across experimental groups. For extracted protein solutions, provide the buffer composition before submission.

Deliverables

The iTRAQ/TMT/MultiNotch Quantitative Proteomics Service typically provides:

1. Protein and Peptide Identification

Identified proteins, peptides, sequence coverage, and supporting MS information.

2. Relative Protein Quantification

Reporter ion-based abundance ratios across labeled samples and experimental groups.

3. Differential Protein Analysis

Statistical comparison of protein abundance changes between predefined groups.

4. Data Visualization

PCA, clustering heatmaps, volcano plots, and other quantitative summary figures.

5. Functional Analysis

GO annotation, KEGG pathway enrichment, and other applicable functional analyses.

6. Raw Data and Final Report

Raw mass spectrometry data, processed result tables, and a structured analysis report.

Start Your Project with MtoZ Biolabs

To discuss whether the iTRAQ/TMT/MultiNotch Quantitative Proteomics Service fits your study, provide the species, sample type, sample number, group design, biological replicates, available protein amount and concentration, planned comparisons, and research objective. MtoZ Biolabs provides a free project consultation to help evaluate service suitability, sample readiness, and the most appropriate quantitative strategy.