MS1-Based Quantification Optimization on DIA Methods on a Quadrupole-Orbitrap Mass Spectrometer

The performance of DIA methods was influenced by the number of DIA windows, the scan resolution, the gradient length and the sampling of the chromatographic peaks. Additionally, the MS1 scan of the DIA method had an important influence on the performance. To optimize quantification on MS1, we evaluated the following conditions. We established a comprehensive spectral library by measuring triplicates on six high pressure reverse phase fractions of Hela and HEK digestion sample. In total, 18 DDA runs are acquired, which results in > 135,000 peptide precursors, > 94,000 peptides, and >7200 protein groups in the spectral library (Table 1). This spectral library is then applied for the DIA data analysis. Figure 1. Data Independent Experiments Design