A DMS Shotgun Lipidomics Workflow Application to Facilitate High-Throughput, Comprehensive Lipidomics

Abstract

Differential mobility spectrometry (DMS) is highly useful for shotgun lipidomic analysis because it overcomes difficulties in measuring isobaric species within a complex lipid sample and allows for acyl tail characterization of phospholipid species. Despite these advantages, the resulting workflow presents technical challenges, including the need to tune the DMS before every batch to update compensative voltages settings within the method. The Sciex Lipidyzer platform uses a Sciex 5500 QTRAP with a DMS (SelexION), an LC system configured for direction infusion experiments, an extensive set of standards designed for quantitative lipidomics, and a software package (Lipidyzer Workflow Manager) that facilitates the workflow and rapidly analyzes the data. Although the Lipidyzer platform remains very useful for DMS-based shotgun lipidomics, the software is no longer updated for current versions of Analyst and Windows. Furthermore, the software is fixed to a single workflow and cannot take advantage of new lipidomics standards or analyze additional lipid species. To address this multitude of issues, we developed Shotgun Lipidomics Assistant (SLA), a Python-based application that facilitates DMS-based lipidomics workflows. SLA provides the user with flexibility in adding and subtracting lipid and standard MRMs. It can report quantitative lipidomics results from raw data in minutes, comparable to the Lipidyzer software. We show that SLA facilitates an expanded lipidomics analysis that measures over 1450 lipid species across 17 (sub)­classes. Lastly, we demonstrate that the SLA performs isotope correction, a feature that was absent from the original software.

Cross-Laboratory Standardization of Preclinical Lipidomics Using Differential Mobility Spectrometry and Multiple Reaction Monitoring

“…At present, the Lipidizer platform misses out on some important lipid classes such as phosphatidylinositol and phosphatidylserine. However, just recently an open access version of the LWM has been published not only enabling workflow adaptations such as the addition of additional classes or the definition and assignment of internal standards but also isotope correction . We believe that this new approach together with the here provided evidence, that standardized methods and protocols lead to reproducible and comparable data, will bring quantitative, comprehensive lipidomics closer to the bedside.…”

“…Consequently, when comparing with previous works, we refrained from isotope correction. Nevertheless, using our recently published algorithm for isotopically correcting the Lipidyzer data, 30 we provide a comparison between corrected and uncorrected values for all investigated NIST materials in Tables S7 and S8 in Supporting Information S2. As can be seen, roughly 85% of all reported lipids present with a deviation of <20%.…”

Divergent MEK/ERK and AMPK signaling dictate lipogenic plasticity and dependence on fatty acid synthesis in Glioblastoma

“…The DMS was tuned with EquiSPLASH LIPIDOMIX (Avanti 330731). Instrument settings, MRM lists, and data analysis methods are available 62 . Quantitative values of lipid species were normalized to cell counts.…”

‘Omic-scale quantitative HILIC-MS/MS approach for circulatory lipid phenotyping in clinical research

“…Advancements in mass spectrometry (MS) technology have improved the sensitivity and selectivity of lipid analysis allowing for the measurement of several hundred lipid species from minimal sample amount in a high-throughput manner 26,27,28 . The most commonly used MS-based approaches for 'omicscale lipid analysis still consist of direct infusion MS techniques (DIMS) using either intra-source 29,30,31 or ion mobility separation (Lipidyzer platform) 32,33,34 . The main drawback of DIMS is the ion suppression and the lack of sensitivity for the quantification of low abundant species [26][27][28][29][30][31][32][33][34][35] .…”

“…The most commonly used MS-based approaches for 'omicscale lipid analysis still consist of direct infusion MS techniques (DIMS) using either intra-source 29,30,31 or ion mobility separation (Lipidyzer platform) 32,33,34 . The main drawback of DIMS is the ion suppression and the lack of sensitivity for the quantification of low abundant species [26][27][28][29][30][31][32][33][34][35] . Therefore, hyphenated techniques with higher separation capacity, such as liquid chromatography (LC) and supercritical fluid chromatography (SFC) coupled to MS, have recently gained significant interest.…”