Systematic Assessment of the Effect of Internal Library in Targeted Analysis of SWATH-MS

Abstract

Targeted analysis of sequential window acquisition of all theoretical massspectra (SWATH-MS) requires the spectral library, which can be generated by shotgun mass spectrometry (MS) or by the pseudo-spectra files directly obtained from SWATHMS data. The external library generated by shotgun MS is employed in most SWATH-MSresearch. However, performance of the internal library, which is constructed by pseudospectra files, in the targeted analysis of SWATH-MS has not been systemically evaluated.Here, we show that up to 40% of the peptides detected by the internal library were not overlapped with those detected by the external library for most SWATH-MS data sets.However, the internal library did not identify extra phosphopeptides compared with the external library for phosphoproteomic SWATH-MS data. Therefore, the internal libraryshould be incorporated into the external library for targeted analysis of nonphosphoproteomic SWATH-MS, given that it can significantly increase the number of peptides of SWATH-MS without requiring additional instrument measurement time Summary. An optional shortened abstract.