Strain-specific peptide (SSP) standard: A genetically encoded quality control standard for isobaric tagging strategies

ABSTRACT: Isobaric tag-based sample multiplexing strategies are extensively used for global protein abundance profiling. However, such analyses are often confounded by ratio compression resulting from the co-isolation, co-fragmentation, and co-quantification of co-eluting peptides, termed “interference.” Recent analytical strategies incorporating ion mobility and real-time database searching have helped to alleviate interference, yet further assessment is needed. Here, we present the Strain-Specific Peptide (SSP) standard, a TMTpro-tagged reference sample that leverages the genetic variation in the proteomes of eight phylogenetically divergent mouse strains. Typically, a peptide with a missense mutation will have a different mass and retention time than the reference or native peptide. TMT reporter ion signal for the native peptide in strains that encode the mutant peptide suggests interference which can be quantified and assessed using the interference-free index (IFI). We showcase the SSP standard by investigating interference in three common data acquisition methods and by testing the improvements in the IFI when using ion mobility-based gas phase fractionation. In addition, we provide a user-friendly, online viewer to visualize the data and streamline calculation of the IFI. The SSP standard will aid in developing and optimizing isobaric tag-based experiments.

INSTRUMENT(S): Orbitrap Eclipse

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Liver

SUBMITTER: Joao Paulo  

LAB HEAD: Joao A. Paulo

PROVIDER: PXD023203 | Pride | 2021-04-03

REPOSITORIES: Pride