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Skyline MS1 Filtering is a label-free quantitative software application that processes full-scan mass spectral data from proteomic experiments by extracting MS1 ion intensity chromatograms across multiple experiments. Key features include comprehensive graphical tools for inspection of MS1 peak picking, visualization of underlying MS/MS spectra, and automated peak integration with manual editing capabilities. Since the introduction of Skyline MS1 Filtering, both instrument vendor support and peptide search pipelines for building spectral libraries have been expanded. New features include run-to-run retention time alignment, time-scheduled import of ion chromatograms (using elution time windows), and facile linking with downstream statistical processing software tools required for high throughput processing of large proteomic discovery datasets. To demonstrate the suitability of MS1 Filtering for quantifying both research and clinical samples, we assessed the workflow reproducibility in monitoring a specific cancer target, receptor tyrosine-protein kinase (ErbB2). We also examined the utility of MS1 Filtering for identifying novel breast cancer biomarker candidates in discovery-based workflows. Taken together, these applications show Skyline MS1 Filtering to be a robust and sensitive approach for high throughput label-free quantification of data-dependent acquisitions. The MS1 Filtering workflow can be seamlessly integrated with selected reaction monitoring assay development within Skyline for further pre-clinical verification.

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