High-resolution mass spectrometry (MS) has become a standard technology for the characterization of proteins, particularly using data-independent acquisition (DIA). The ability to detect and quantify proteins across a wide dynamic range requires achieving both high sensitivity and maximum depth of proteome coverage. For this purpose, nanoflow liquid chromatography (LC) is often used. Maximizing MS performance can be realized through improvements in nanoflow chromatographic separation, specifically with narrower LC peak shapes and broader separation of complex mixtures of peptides.
This poster summarizes the use of Zeno SWATH DIA on the SCIEX ZenoTOF 7600 system in combination with nanoflow chromatographic separation using IonOpticks Aurora SX series columns. Maximum sensitivity is demonstrated with the detection of >2,300 protein groups (>10,000 precursors) in low column loads (250 pg) of a complex digest standard, approximating single-cell protein levels. To show the depth of proteome coverage and quantitative fidelity, mixtures of digests from 3 separate organisms (human, yeast and E. coli) at different ratios were analyzed, whereby >19,000 protein groups were detected in the mixtures with >94% showing quantitative reproducibility of <20% CV and protein ratios between different mixtures accurately quantitated using a label-free quantitation (LFQ) strategy with Zeno SWATH DIA.