mIMT-visHTS: A novel method for multiplexing isobaric mass tagged datasets with an accompanying visualization high throughput screening tool for protein profiling

Abstract Isobaric mass tagging (IMT) methods enable the analysis of thousands of proteins simultaneously. We used tandem mass tagging reagents (TMT™) to monitor the relative changes in the proteome of the mouse macrophage cell line RAW264.7 at the same six time points after no stimulation (baseline phenotype), stimulation with interferon gamma (pro-inflammatory phenotype) or stimulation with interleukin-4 (anti-inflammatory phenotype). The combined TMT datasets yielded nearly 12,000 protein profiles for comparison. To facilitate this large analysis, we developed a novel method that combines or multiplexes the separate IMT (mIMT) datasets into a single super dataset for subsequent model-based clustering and co-regulation analysis. Specially designed visual High Throughput Screening (visHTS) software screened co-regulated proteins. visHTS generates an interactive and visually intuitive color-coded bullseye plot that enables users to browse the cluster outputs and identify co-regulated proteins.