A Comprehensive Proteome of Human Corneal Epithelial Cells Constructed by Cross-platform DIA-Mass Spectrometry.

The corneal epithelium serves as the front barrier against environmental stimuli and pathogens on the ocular surface. A comprehensive protein profile of the corneal epithelium would be crucial for understanding the molecular mechanisms that are related to corneal disease. This work demonstrated a library-free data-independent acquisition (DIA) approach across different mass spectrometers and proteomic software to build a comprehensive proteomic dataset for human corneal epithelial cells (HCECs). With the combinational use of different data-independent acquisition technologies of multiple mass spectrometers, including Sciex ZenoTOF 7600 (DIA-SWATH), Bruker TimsTOF Pro2 (DIA-PASEF), and ThermoFisher Orbitrap Fusion Lumos (DIA-HRMS1), protein identification and quantification were performed with superior sensitivity and resolution. By using a library-free DIA approach, this study constructed a more diverse and unbiased proteomic profile of human corneal epithelial cells (HCECs), comprising 11,954 protein groups (1% FDR). This represents the largest corneal proteome reported to date. All raw proteomic data were deposited to ProteomeXchange Consortium via Proteomics Identifications database (PRIDE) with the dataset identifier accession number PXD059451. Our findings hold the potential to enhance future understanding of corneal pathologies and transformative therapeutics.