Maximizing Flexibility on a High Resolution Open Access LC/MS System

Theresa McLaughlin, Allis S. Chien

Stanford University Mass Spectrometry, Stanford, CA

First Published: ABRF, Palm Springs, CA, Poster 188, March 2, 2020.

Introduction

Traditionally, mass spectrometers commonly available to open access core facility users are low resolution GC-EIMS or LC-ESIMS with single quadrupole mass filters. These instruments are robust and very useful for small molecule synthetic chemists. However, with advances in the fields of metabolomics and biotherapeutics there is a need for on-demand methods supporting a wider range of analytes, from amino acids to intact proteins. High resolution time of flight and orbitrap mass spectrometers are the instruments of choice for intact protein analysis and support many qualitative and quantitative small molecule workflows. They have the mass stability and operational reliability needed for an open access lab setting. Our challenge is to configure a single LC-ESIHRMS system to support a broad spectrum of users.

Instrument Methods

Our LC-ESIHRMS system is a Waters Acquity H Class UPLC with a Thermo Exactive Orbitrap mass spectrometer. The Acquity includes an automated switching valve for 4 columns plus a bypass for Direct Injection methods. The Exactive is mass calibrated once a week and maintains 5 ppm mass accuracy with no internal standard.

The system includes columns to support a variety of sample types: C-18 for small molecules, C-8 for peptides and small proteins, Polyphenyl for intact proteins, and HILIC for small, polar molecules. Four solvents are available: 0.1% formic acid in water (A), 0.1% formic acid in acetonitrile (B), 5 mM ammonium acetate in 0.1% formic acid
(C), and methanol (D). Typical examples are shown in Figures 1 to 4.

RemoteAnalyzer software from SpectralWorks simplifies the user experience. It allows users to input their sample information and choose from a menu of pre-built instrument methods, generates reports, and creates a link to the raw data when the run is complete. The software supports multiple instrument combinations
(Agilent, Bruker, Thermo, Waters, etc.).

Discussion

The flexibility of this open access LC-ESIHRMS setup has improved turnaround time for a variety of sample types. Clients who take advantage of the open access workflow can submit their own samples and have their results the same day. Laboratory staff are also able to work more efficiently, as switching on the fly among reversed phase, HILIC, and intact protein methods enables staff to queue and analyze user-submitted samples upon receipt, without manual reconfiguration between sample types. The typical Open Access Workflow is shown in Figure 5.

Fig 5. Open Access Workflow.

Open access users assume increased responsibility for experimental design and data processing. Initial training emphasizes the importance of running blanks and system suitability tests, as well as positive and negative controls during sample analysis. Some projects, such as those involving trace analysis of precious samples, may be better served by an instrument with controlled access.

The RemoteAnalyzer software is intuitive and enables users to remotely view instrument status, queue length, and to access their results via a web browser. For data analysis, in addition to Thermo’s Xcalibur software, vendor-neutral tools are available including the SpectralWorks browser, MestReNova, and ProteinMetrics for deconvolution of intact protein spectra.

Acknowledgements

This work is supported by the Stanford Dean of Research and the Vincent and Stella Coates Foundation.

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