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High-resolution enabled 12-plex DiLeu isobaric tags for quantitative proteomics.

Dustin C Frost1, Tyler Greer, Lingjun Li

  • 1School of Pharmacy, University of Wisconsin , 777 Highland Avenue, Madison, Wisconsin 53705, United States.

Analytical Chemistry
|November 19, 2014
PubMed
Summary
This summary is machine-generated.

Researchers developed DiLeu, a cost-effective multiplex isobaric tag for mass spectrometry. This new reagent enables 12-plex quantitative proteomics, enhancing throughput and reducing costs in proteomic analyses.

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Area of Science:

  • Proteomics
  • Analytical Chemistry
  • Biochemistry

Background:

  • Multiplex isobaric tags are essential for high-throughput quantitative proteomics using mass spectrometry.
  • Commercial tags like TMT and iTRAQ offer valuable quantitative capabilities but can be expensive.
  • In-house synthesis of proteomic reagents offers a cost-effective alternative for researchers.

Purpose of the Study:

  • To develop a cost-effective, in-house synthesized multiplex isobaric tag for quantitative proteomics.
  • To increase the multiplexing capacity of the DiLeu reagent beyond existing capabilities.
  • To maintain or improve the quantitative performance of the DiLeu reagent.

Main Methods:

  • Development of novel DiLeu multiplex isobaric tags utilizing selective incorporation of stable isotopes ((13)C, (15)N, (2)H).
  • Exploitation of mass defects to create new reporter isotopologues with precise mass differences (6 mDa).
  • High-resolution nano liquid chromatography-tandem mass spectrometry (nanoLC-MS(2)) analysis on an Orbitrap Elite mass spectrometer.

Main Results:

  • Achieved a 3-fold increase in multiplexing capacity, creating a 12-plex isobaric set.
  • Demonstrated baseline resolution of new reporter variants in high-resolution HCD spectra.
  • Showcased accurate 12-plex quantitation of Saccharomyces cerevisiae lysate digest using the enhanced DiLeu tags.

Conclusions:

  • The enhanced DiLeu reagent provides a cost-effective, high-performance alternative for 12-plex quantitative proteomics.
  • The synthetic simplicity and quantitative accuracy are preserved with the expanded multiplexing capacity.
  • This advancement facilitates more comprehensive and economical proteomic studies.