Poster Presentation HUPO 2019 - 18th Human Proteome Organization World Congress

Towards Turnkey Targeted Proteomics Solutions Using Internal Standard Triggered Acquisitions on Next Generation Orbitrap Mass Spectrometers (#723)

Sebastien Gallien 1 2 , Aaron S Gajadhar 3 , Bhavin Patel 4 , Markus Kellmann 5 , Tabiwang N Arrey 5 , Christian Thoeing 5 , Alexander Harder 5 , Romain Huguet 3 , Graeme McAlister 3 , Derek Bailey 3 , Shannon Eliuk 3 , Emily I Chen 2 , Yue Xuan 5 , Andreas Huhmer 3 , Ken Miller 3
  1. Thermo Fisher Scientific, Paris, France
  2. Thermo Fisher Scientific, Precision Medicine Science Center, Cambridge, MA, USA
  3. Thermo Fisher Scientific, San Jose, CA, USA
  4. Thermo Fisher Scientific, Rockford, IL, USA
  5. Thermo Fisher Scientific, Bremen, Germany

Targeted quantitative proteomics based on high resolution parallel reaction monitoring (PRM) technique benefits from improved measurement selectivity, allowing more sensitive endogenous peptides quantification in complex samples. Here we introduce an extension of PRM, called “SureQuant” method, which uses spiked-in internal standards (IS) to dynamically control the acquisition process and to maximize its productivity. This novel method has been implemented in the native instrument control software of next-generation Orbitrap instruments, Thermo Scientific™ Orbitrap Exploris™ 480 and Eclipse™ Tribrid™ mass spectrometers, to enable a broad access. The SureQuant IS targeted protein quantification method has been adapted from the IS-PRM method in order to improve its usability and robustness (especially against chromatographic variations). Its ability to deliver high-density, ultra-sensitive measurements has benefited to a variety of applications. Applied to the monitoring of signaling pathways in cell lines and tissues specimens supplemented with stable isotopically labeled (SIL) peptides (30-150 IS, including Thermo Scientific™ Pierce™ SureQuant™ kits), the method enabled systematic quantification of endogenous peptides with high precision (<5%-CV for the majority of peptides) and short analysis time (10-40 min LC gradient). In a larger scale application of the method, non-depleted plasma samples supplemented with 804 SIL peptides (Biognosys™ PQ500™ kit) were analyzed with a 70-min LC gradient for global plasma proteome quantification. More than 550 endogenous peptides, surrogates of around 400 plasma proteins, were reproducibly quantified over a 6 orders of magnitude range. This proteome coverage compared favorably with that of profiling methods, while still benefiting from the enhanced data quality of targeted measurements (including peptide quantification in the low amol range). The multiple analytical benefits of the SureQuant method combined with the ability to embed pre-set (optimized) methods, associated with predefined kits of IS peptides, directly into the instrument control software represents a decisive step towards the provision of turnkey targeted proteomics solutions.