title: Versatile, sensitive liquid chromatography mass spectrometry - Implementation of 10 μm OT columns suitable for small molecules, peptides and proteins.
abstract: We have designed a versatile and sensitive liquid chromatographic (LC) system, featuring a monolithic trap column and a very narrow (10 μm ID) fused silica open tubular liquid chromatography (OTLC) separation column functionalized with C18-groups, for separating a wide range of molecules (from small metabolites to intact proteins). Compared to today's capillary/nanoLC approaches, our system provides significantly enhanced sensitivity (up to several orders) with matching or improved separation efficiency, and highly repeatable chromatographic performance. The chemical properties of the trap column and the analytical column were fine-tuned to obtain practical sample loading capacities (above 2 μg), an earlier bottleneck of OTLC. Using the OTLC system (combined with Orbitrap mass spectrometry), we could perform targeted metabolomics of sub-μg amounts of exosomes with 25 attogram detection limit of a breast cancer-related hydroxylated cholesterol. With the same set-up, sensitive bottom-up proteomics (targeted and untargeted) was possible, and high-resolving intact protein analysis. In contrast to state-of-the-art packed columns, our platform performs chromatography with very little dilution and is "fit-for-all", well suited for comprehensive analysis of limited samples, and has potential as a tool for challenges in diagnostics.
authors.affiliation: Department of Chemistry, University of Oslo, Post Box 1033 Blindern, NO-0315 Oslo, Norway.|Department of Engineering Sciences, University of Agder, Jon Lilletunsvei 9, NO-4891 Grimstad, Norway.
authors.forename: T
authors.initials: T
authors.lastname: Vehus
authors.affiliation: Department of Chemistry, University of Oslo, Post Box 1033 Blindern, NO-0315 Oslo, Norway.
authors.forename: H
authors.initials: H
authors.lastname: Roberg-Larsen
authors.affiliation: Unit for Cell Signaling, SFI-CAST Biomedical Innovation Center, Oslo University Hospital, Rikshospitalet, NO-0027 Oslo, Norway.
authors.forename: J
authors.initials: J
authors.lastname: Waaler
authors.affiliation: Unit for Cell Signaling, SFI-CAST Biomedical Innovation Center, Oslo University Hospital, Rikshospitalet, NO-0027 Oslo, Norway.
authors.forename: S
authors.initials: S
authors.lastname: Aslaksen
authors.affiliation: Unit for Cell Signaling, SFI-CAST Biomedical Innovation Center, Oslo University Hospital, Rikshospitalet, NO-0027 Oslo, Norway.
authors.forename: S
authors.initials: S
authors.lastname: Krauss
authors.affiliation: Department of Chemistry, University of Oslo, Post Box 1033 Blindern, NO-0315 Oslo, Norway.
authors.forename: S R
authors.initials: SR
authors.lastname: Wilson
authors.affiliation: Department of Chemistry, University of Oslo, Post Box 1033 Blindern, NO-0315 Oslo, Norway.
authors.forename: E
authors.initials: E
authors.lastname: Lundanes
mesh_terms: D000818:Animals; D060466:Axin Protein / Q000302:isolation & purification; D001943:Breast Neoplasms / Q000737:chemistry / Q000175:diagnosis* / Q000378:metabolism; D002853:Chromatography, Liquid / Q000295:instrumentation* / Q000379:methods; D055354:Exosomes / Q000737:chemistry; D005260:Female; D006801:Humans; D006888:Hydroxycholesterols / Q000302:isolation & purification*; D051379:Mice; D010455:Peptides / Q000302:isolation & purification*; D040901:Proteomics / Q000295:instrumentation* / Q000379:methods; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry / Q000295:instrumentation* / Q000379:methods
chemical_list: D060466:Axin Protein; C555452:Axin1 protein, mouse; D006888:Hydroxycholesterols; D010455:Peptides