A Triple KnockOut Isobaric-labelling Quality Control Platform in an Integrated On-line Database Search

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 co-eluting, 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 Knock-Out (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.

采用等压标记（isobaric tagging）的样本多重分析是实现全蛋白质组蛋白定量的高效策略。等压标记技术的一大主要局限为比值压缩现象：该现象由共洗脱的近等压肽段的分离、碎裂与定量过程引发，通常被称为“离子干扰（ion interference）”。随着新型仪器与分析方法不断迭代升级，搭建一款可实现质量控制、参数优化并支持跨样本比较的稳健平台已成为必要需求。本研究介绍了TKO-iQC这一集成化平台，其由三重敲除（Triple Knock-Out, TKO）酵母酶解标准品，以及一款自动化网页端数据库检索与蛋白谱可视化应用程序共同构成。我们还展示了两款基于TMTpro试剂的新型TKO标准品（TKOpro9与TKOpro16），以及一款升级后的TKO查看工具TVT2.0。TKO-iQC凭借简洁流畅的工作流程，极大简化了仪器性能的对比工作。