Author:
Lin Weiwei,Mousavi Fatemeh,Blum Benjamin C.,Heckendorf Christian F.,Lampl Noah,Heckman Ryan,Guo Hongbo,McComb Mark,Emili Andrew
Abstract
SummaryHigh sensitivity nanoflow liquid chromatography (nLC) is seldom employed in untargeted metabolomics because current sample preparation techniques are inefficient to prevent nanocapillary column performance degradation. Here, we describe an nLC-based tandem mass spectrometry workflow that enables seamless joint analysis and integration of metabolomics (including lipidomics) and proteomics from the same samples without instrument duplication. This workflow is based on robust solid phase micro-extraction step for routine sample clean-up and bioactive molecule enrichment. Our method, termed PANAMA, improves compound resolution and detection sensitivity without compromising depth of coverage as compared with existing widely used analytical procedures. Notably, PANAMA can be applied to a broad array of specimens including biofluids, cell line and tissue samples. It generates high quality, information rich metabolite-protein datasets while bypassing the need for specialized instrumentation.MotivationThe ability to routinely, sensitively and reproducibly analyze both cellular proteins and metabolite mixtures from the same biospecimens can enhance the discovery of biomolecules associated with basic biochemical processes and pathobiological states. Yet existing mass spectrometry-based profiling methods rely on specialized protocols and duplicated instrumentation platforms, resulting in increased time, sample consumption and costs. We sought to generate an effective platform for both metabolomic and proteomic studies on the same samples by enabling nanoflow liquid chromatography for small molecules. The resulting approach was extensively optimized and benchmarked to provide in depth molecular coverage, along with improved chromatographic separations, sensitivity and reliability as compared to existing methods. The cost benefit ratio of PANAMA is substantial because the platform bypasses the need for specialized instrumentation stemming from incompatible procedures.
Publisher
Cold Spring Harbor Laboratory