Label-Free Quantification from Direct Infusion Shotgun Proteome Analysis (DISPA-LFQ) with 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 an improved direct infusion shotgun proteome
analysis protocol including label-free quantification (DISPA-LFQ)
using CsoDIAq software. With CsoDIAq analysis of DISPA data, we can
now identify up to ∼2000 proteins from the HeLa and 293T proteomes,
and with DISPA-LFQ, we can quantify ∼1000 proteins from no
more than 1 μg of sample within minutes. The identified proteins
are involved in numerous valuable pathways including central carbon
metabolism, nucleic acid replication and transport, protein synthesis,
and endocytosis. Together with a high-throughput sample preparation
method in a 96-well plate, we further demonstrate the utility of this
technology for performing high-throughput drug analysis in human 293T
cells. The total time for data collection from a whole 96-well plate
is approximately 8 h. We conclude that the DISPA-LFQ strategy presents
a valuable tool for fast identification and quantification of proteins
in complex mixtures, which will power a high-throughput proteomic
era of drug screening, biomarker discovery, and clinical analysis.