Extracellular Vesicle Analysis by Paper Spray Ionization Mass Spectrometry

Abstract

Paper spray ionization mass spectrometry (PSI-MS) is a direct MS analysis technique with several reported bacterial metabolomics applications. As with most MS-based bacterial studies, all currently reported PSI-MS bacterial analyses have focused on the chemical signatures of the cellular unit. One dimension of the bacterial metabolome that is often lost in such analyses is the exometabolome (extracellular metabolome), including secreted metabolites, lipids, and peptides. A key component of the bacterial exometabolome that is gaining increased attention in the microbiology and biomedical communities is extracellular vesicles (EVs). These excreted structures, produced by cells in all domains of life, contain a variety of biomolecules responsible for a wide array of cellular functions, thus representing a core component of the bacterial secreted metabolome. Although previously examined using other MS approaches, no reports currently exist for a PSI-MS analysis of bacterial EVs, nor EVs from any other organism (exosomes, ectosomes, etc.). PSI-MS holds unique analytical strengths over other commonly used MS platforms and could thus provide an advantageous approach to EV metabolomics. To address this, we report a novel application representing, to our knowledge, the first PSI-MS analysis of EVs from any organism (using the human gut resident Oxalobacter formigenes as the experimental model, a bacterium whose EVs were never previously investigated). In this report, we show how we isolated and purified EVs from bacterial culture supernatant by EV-specific affinity chromatography, confirmed and characterized these vesicles by nanoparticle tracking analysis, analyzed the EV isolate by PSI-MS, and identified a panel of EV-derived metabolites, lipids, and peptides. This work serves as a pioneering study in the field of MS-based EV analysis and provides a new, rapid, sensitive, and economical approach to EV metabolomics.

Data-Driven Soft Independent Modeling of Class Analogy in Paper Spray Ionization Mass Spectrometry-Based Metabolomics for Rapid Detection of Prostate Cancer

“…PSI-HRMS mass spectra were obtained using a Thermo Scientific Q Exactive MS. Data acquisition was performed in full scan mode (positive ionization mode) with a mass range of 70–1,000 m / z at 140,000 mass resolution. Other PSI-HRMS instrument parameters for data acquisition were used as previously described. ,, Metabolomics data files were converted to mzXML file format using RawConverter . Mzmine 2.0 was employed for peak picking and feature alignment .…”

“…Other PSI-HRMS instrument parameters for data acquisition were used as previously described. 5,39,40 Metabolomics data files were converted to mzXML file format using RawConverter. 41 Mzmine 2.0 was employed for peak picking and feature alignment.…”

Embossed Paper Platform for Whole Blood Collection, Room Temperature Storage, and Direct Analysis by Pinhole Paper Spray Mass Spectrometry

“…This approach is different from all other microsampling techniques such as capillary microsampling, volumetric absorptive microsampling, plasma card collections, Schirmer paper strip for sampling 1 μL of tear fluid, and dried blood spots (DBS) that either sample liquid blood or absorb blood into a hydrophilic cellulose substrate. Due to their ability to collect a smaller volume of blood samples (<50 μL), microsampling platforms have been widely employed in a large range of applications, from pharmacokinetic or toxicokinetic studies, , clinical diagnostics (e.g., newborn screening programs), − to therapeutic drug monitoring. − Altogether, microsampling platforms can reduce the need for a centralized laboratory model, making personalized healthcare more widely accessible to individuals allowing for faster turnaround time for testing and treatment.…”

Applications of ambient ionization mass spectrometry in 2021: An annual review

“…In addition to the application of AIMS to tissue imaging and biological fluids, recent applications of AIMS also include the study of protein folding and solubility, 134,135 the quantification of extracellular neurotransmitters in brain tissue 136 and the characterization of metabolites in bacterial extracellular vesicles (EVs). 137 The ability to perform real-time analysis of bacteria headspace has also been achieved using AIMS. In medical diagnostics, the ability to quickly identify bacterial species is crucial in ensuring the appropri-…”