Fig. 2.

Averaged MS1 precursorlibrary matching (AMPL) increases peptide detection sensitivity.A, schematic outlining the AMPL experimental design. B, both the AMPL and BoxCar acquisition methods prioritize MS time to enhance MS1 scan quality. Schematic comparing duty cycles for data-dependent acquisition (DDA), AMPL, and BoxCar acquisition methods on the indicated MS instruments (Orbitrap Elite, Orbitrap HF). The median peak width using our chromatographic setup with the Orbitrap Elite is ∼38 s. C, a comparison between AMPL and DDA + L, showing intensity distributions of peptide features identified by MS/MS (blue) and matching to identified library features (red). D, schematic outlining experimental workflow for assessing match-between-runs false discovery rate. E, features matched in target and decoy proteomes before and after additional filtering based on match retention time difference, match m/z difference, and match m/z error. F and G, features (F) and unique peptides (G) detected in AMP(L) versus DDA. DDA + L is DDA with matching to a library. H, proteome coverage versus cell number. The cell titration was performed in duplicate.