Figure 1.

An overview of data acquisition and spectral library generation workflow. Data acquisition: Pooled cervico-vaginal fluid samples from 9 cancer cases, 3 AH and 7 controls were used for the library generation while two EC cell lines were used for the cell line-based library. Cervico-vaginal samples were separated into pellets and supernatants. Pellets from both cervico-vaginal fluid and EC cell lines were subjected to lysis and protein extraction. All samples were subjected to proteolytic digestion with trypsin, peptide fractionation using high-pH reverse phase liquid chromatography and LC-MS/MS analysis. Library generation: Sequence database search was done using X!Tandem while the library construction was performed using the OpenSWATH pipeline. Library validation: technical replicates of five cervico-vaginal samples were used to search the library to assess its applicability for SWATH-MS data analysis, while ten samples (five cancers and five controls) were used to assess the potential utility of the library for biomarker discovery in the real-world setting.