Enhanced sample multiplexing-based targeted proteomics with intelligent data acquisition

ABSTRACT: Targeted proteomics has been playing an increasingly important role in hypothesis-driven protein research and clinical bi-omarker discovery. We have previously created a workflow, Tomahto, to enable real-time targeted pathway proteomics assays using two-dimensional multiplexing technology. Coupled with the TMT 11-plex reagent, hundreds of proteins of in-terest from up to 11 samples can be targeted and accurately quantified in a single-shot experiment with remarkable sensi-tive. However, room remains to further improve sensitivity, accuracy, and throughput, especially for targeted studies de-manding a high peptide-level success rate. Here, bearing in mind the goal to improve peptide-level targeting, we introduce several new functionalities in Tomahto, featuring integration of gas-phase fractionation using the FAIMS device, an ac-companying software program (TomahtoPrimer) to customize fragmentation for each peptide target, and support for higher multiplexing capacity with the latest TMTpro18-plex reagent. We demonstrate that adding these features to the Tomahto platform significantly improves overall success rate from 89% to 98% in a single 60-min targeted assay of 290 peptides across human cell lines, while boosting quantitative accuracy via reducing TMT reporter ion interference.

INSTRUMENT(S): Orbitrap Eclipse

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Qing Yu  

PROVIDER: MSV000095604 | MassIVE | Wed Aug 14 14:02:00 BST 2024

REPOSITORIES: MassIVE