Rapid Profiling of 2000 Human Protein Groups with Direct Infusion Shotgun Proteome Analysis (DISPA) + CsoDIAq Software

Large-scale proteome analysis requires rapid and high-throughput analytical methods. We recently reported a new paradigm in proteome analysis where direct infusion and ion mobility are used instead of liquid chromatography (LC) to achieve rapid and high-throughput proteome analysis. Here we introduce several improvements to direct infusion shotgun proteome analysis (DISPA) to achieve identification of over 2,000 human protein groups, which is nearly four-fold more than the original publication. With DISPA using an Orbitrap Exploris 240 DISPA + CsoDIAq data analysis software, we demonstrate a rapid identification of over 2000 proteins from the HeLa proteome and accurate label free quantification (LFQ) of 1045 proteins from no more than one microgram of sample within minutes. From each single spectrum multiple peptides are often identified, up to 36 in one case. The identified proteins are involved in numerous valuable pathways including central carbon metabolism, nucleic acid replication and transport, protein synthesis, and endocytosis. We conclude that the DISPA + CsoDIAq strategy presents a valuable tool for high-throughput proteome analysis, enabling fast identification and quantification of proteins in complex mixtures.