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Related Concept Videos

Peptide Identification Using Tandem Mass Spectrometry01:33

Peptide Identification Using Tandem Mass Spectrometry

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Tandem mass spectrometry, also known as MS/MS or MS2, is an analytical technique that employs two mass analyzers. Essentially it is a series of mass spectrometers that helps isolate a particular biomolecule and then helps study its chemical properties.
This technique helps gather information regarding the protein from which the peptide was obtained and to study the peptides’ amino acid sequence. Identifying peptides from a complex mixture is an important component of the growing field of...
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TMT Sample Preparation for Proteomics Facility Submission and Subsequent Data Analysis
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Automated Sample Preparation Workflow for Tandem Mass Tag-Based Proteomics.

Dong-Gi Mun1, Neha S Joshi1,2, Rohit Budhraja1

  • 1Department of Laboratory Medicine and Pathology, Mayo Clinic, 200 First Street SW, Rochester, Minnesota 55905, United States.

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|September 1, 2023
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Summary

Automating tandem mass tag (TMT) labeling for protein quantitation using the AccelerOme platform matches manual methods in efficiency and accuracy. This automated workflow offers robust results and high throughput for large-scale TMT-based proteomic studies.

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Area of Science:

  • Proteomics
  • Mass Spectrometry
  • Biotechnology

Background:

  • Tandem mass tag (TMT)-based isobaric labeling is crucial for multiplexed protein quantitation.
  • Manual TMT labeling workflows are complex, requiring high proficiency and introducing variability, hindering widespread adoption, especially in large-scale studies.

Purpose of the Study:

  • To compare the performance of the AccelerOme automated platform for TMT labeling with conventional manual sample processing.
  • To evaluate the efficiency, accuracy, and reproducibility of automated TMT-based proteomics.

Main Methods:

  • A 16-plex TMT experiment was conducted using both the AccelerOme automated platform and a manual protocol on cell pellets.
  • Samples were analyzed using single-shot liquid chromatography-tandem mass spectrometry (LC-MS/MS).

Main Results:

  • The automated platform identified a higher number of proteins and peptides compared to the manual method.
  • Tryptic digestion, alkylation, and TMT labeling efficiencies were comparable between automated and manual processes.
  • Quantitation accuracy and precision were similar for both workflows, with the automated platform performing on par with expert manual preparation.

Conclusions:

  • The AccelerOme automated platform demonstrates comparable performance to expert manual sample preparation for TMT-based proteomics.
  • This automated workflow enhances reproducibility and throughput, making it suitable for large-scale TMT-based studies.
  • The platform has the potential to replace manual pipelines, improving efficiency and robustness in proteomic quantitation.