Quantitative Analysis of Targeted Proteins in Complex Sample Using Novel Data Independent Acquisition

Abstract

Abstract Data Independent Acquisition (DIA) is a novel mass spectrometry (MS) scan mode for quantitative proteomics, acquiring all precursors as well as fragments without losing the low abundant ions. This method overcome the limitation of product ion quantification by high-resolution MS. We developed three DIA methods on Q-qIT-OT Tribrid MS, classic DIA, novel Wide isolation window SIM scan DIA (WiSIM-DIA) and Full MS-DIA. WiSIM-DIA utilizes ultra-high resolution SIM scan for quantification, complementary with classic DIA. Quantitative analysis of 10 low abundant peptides spiked in Hela digest was performed by these three methods for linearity, reproducibility and sensitivity evaluation. The results showed that the LOQs reached amol level (14−435 amol) with good linearity and effective MS/MS confirmation. The isolation window of Full MS-DIA was as low as 3 amu, and the data can be directly used for database searching. This method realized the integration of Data dependent acquisition (DDA) and DIA, and thus overcoming the limitation of using spectra library.