Proteomic data acquirement and analysis
HF mass spectrometer, PCOS
SEPARATION
Plasma protein mass spectrometry analysis was conducted in 3 patients in each group. Specifically, 20 μg protein was separated into peptide fractions by an LC-20AB HPLC system (Shimadzu, Kyoto, Japan). Peptide fractions were loaded onto a Thermo UltiMate 3000 UHPLC system coupled with a Q-Exactive HF mass spectrometer (Thermo Fisher Scientific, San Jose, CA, USA) for peptide separation and identification. A proteome spectral library was built in data-dependent acquisition (DDA) mode, followed by peptide identification in data-independent acquisition (DIA) mode using a sequential window acquisition of all theoretical fragments (SWATH) method. In DDA mode, the top 20 precursors with charge statuses ranging from +2 to +6 were selected for fragmentation during the MS2 scan. For spectral data analysis, peptide identification was conducted using MaxQuant ver. 1.5.3.30 against the UniProtKB/Swiss-Prot protein knowledgebase (Release 2019_06, Proteomic analysis of plasma from PCOS pa tients treated with CPA/EE+Met or GLP-1RA+Met. Volcano plots of differentially expressed plasma proteins from patients prior to or after treatment with CPA/EE+Met (