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Sample multiplexing using isobaric tagging is a powerful strategy for proteome-wide protein quantification. One major caveat of isobaric tagging is ratio compression that results from the isolation, fragmentation, and quantification of coeluting, near-isobaric peptides, a phenomenon typically referred to as "ion interference". A robust platform to ensure quality control, optimize parameters, and enable comparisons across samples is essential as new instrumentation and analytical methods evolve. Here, we introduce TKO-iQC, an integrated platform consisting of the Triple Knockout (TKO) yeast digest standard and an automated web-based database search and protein profile visualization application. We highlight two new TKO standards based on the TMTpro reagent (TKOpro9 and TKOpro16) as well as an updated TKO Viewing Tool, TVT2.0. TKO-iQC greatly facilitates the comparison of instrument performance with a straightforward and streamlined workflow.

A Triple Knockout Isobaric-Labeling Quality Control Platform with an Integrated Online Database Search