Oral Presentation 24th Annual Lorne Proteomics Symposium 2019

Evaluation of a novel LC system that embeds analytes in pre-formed gradients for rapid, ultra-robust proteomics (#53)

Nicolai Bache 1 , Philipp Geyer 2 , Dorte Bekker-Jensen 3 , Ole Hoerning 1 , Lasse Falkenby 1 , Peter Treit 2 , Sophia Doll 2 , Igor Paron 2 , Florian Meier 2 , Jesper Olsen 3 , Ole Vorm 1 , Matthias Mann 2
  1. Evosep Biosystems, Odense, FYN, Denmark
  2. Department of Proteomics and Signal Transduction, Max Planck Institute of Biochemistry, Martinsried, Bavaria, Germany
  3. Novo Nordisk Foundation Center for Protein Research, Proteomics Program, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Seeland, Denmark

BackgroundMass spectrometry-based proteomics and metabolomics are fast growing and powerful technologies, with the potential to revolutionise health care and precision medicine. However, available separation systems have so far limited throughput and robustness and thereby prevented omic technologies from being fully integrated and routinely applied in clinical settings. Here, we evaluate a conceptually novel liquid chromatography (LC) system that significantly increases robustness and sample throughput while maintaining the sensitivity of current nano-flow LC. MethodsThe new system, called Evosep One, uses four low-pressure pumps in parallel to elute samples from a disposable and single use trap column while also forming a chromatographic gradient. The sample and gradient are moved into a storage loop that subsequently is switched in-line with a single high-pressure pump and an analytical separation column for separation. ResultsWe have characterised the performance of the new system regarding cross contaminations (<0.07%), retention time shifts and peak width (<3 sec) in over 1500 HeLa runs. The short overhead time of approximately 2 min allows us to efficiently measure 300, 200, 100, 60 or 30 samples per day with corresponding gradient lengths of 3.2, 5.6, 11.5, 21 and 44 minutes, respectively. The performance and applicability in various proteomics LC-MS strategies was evaluated for simple, medium and complex sample types. From fractionated HeLa cell lysates, deep proteomes covering more than 130,000 sequence unique peptides and around 10,000 proteins were rapidly acquired (18 h total instrument time). Using this data as a library for data independent acquisition, we demonstrate the quantitation of 5200 proteins in only 21 min. ConclusionWe evaluated and benchmarked how to use the Evosep One in cutting edge LC-MS strategies to significantly increase overall performance and throughput. We also demonstrate how this can be applied to clinical research workflows that require uninterrupted analysis of thousands of crude biological samples as well as routine applications such as doping control and drug screening.