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Analysis of a common cold virus and its subviral particles by gas-phase electrophoretic mobility molecular analysis and native mass spectrometry.

ABSTRACT: Gas-phase electrophoretic mobility molecular analysis (GEMMA) separates nanometer-sized, single-charged particles according to their electrophoretic mobility (EM) diameter after transition to the gas-phase via a nano electrospray process. Electrospraying as a soft desorption/ionization technique preserves noncovalent biospecific interactions. GEMMA is therefore well suited for the analysis of intact viruses and subviral particles targeting questions related to particle size, bioaffinity, and purity of preparations. By correlating the EM diameter to the molecular mass (Mr) of standards, the Mr of analytes can be determined. Here, we demonstrate (i) the use of GEMMA in purity assessment of a preparation of a common cold virus (human rhinovirus serotype 2, HRV-A2) and (ii) the analysis of subviral HRV-A2 particles derived from such a preparation. (iii) Likewise, native mass spectrometry was employed to obtain spectra of intact HRV-A2 virions and empty viral capsids (B-particles). Charge state resolution for the latter allowed its Mr determination. (iv) Cumulatively, the data measured and published earlier were used to establish a correlation between the Mr and EM diameter for a range of globular proteins and the intact virions. Although a good correlation resulted from this analysis, we noticed a discrepancy especially for the empty and subviral particles. This demonstrates the influence of genome encapsulation (preventing analytes from shrinking upon transition into the gas-phase) on the measured analyte EM diameter. To conclude, GEMMA is useful for the determination of the Mr of intact viruses but needs to be employed with caution when subviral particles or even empty viral capsids are targeted. The latter could be analyzed by native MS.

SUBMITTER: Weiss VU

PROVIDER: S-EPMC4558612 | biostudies-literature | 2015 Sep

REPOSITORIES: biostudies-literature

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Analysis of a common cold virus and its subviral particles by gas-phase electrophoretic mobility molecular analysis and native mass spectrometry.

Weiss Victor U VU Bereszcazk Jessica Z JZ Havlik Marlene M Kallinger Peter P Gösler Irene I Kumar Mohit M Blaas Dieter D Marchetti-Deschmann Martina M Heck Albert J R AJ Szymanski Wladyslaw W WW Allmaier Günter G

Analytical chemistry 20150812 17

Gas-phase electrophoretic mobility molecular analysis (GEMMA) separates nanometer-sized, single-charged particles according to their electrophoretic mobility (EM) diameter after transition to the gas-phase via a nano electrospray process. Electrospraying as a soft desorption/ionization technique preserves noncovalent biospecific interactions. GEMMA is therefore well suited for the analysis of intact viruses and subviral particles targeting questions related to particle size, bioaffinity, and pur ...[more]

PMID: 26221912

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Project description:The emerging role of extracellular vesicles (EVs) as biomarkers and their envisioned therapeutic use require advanced techniques for their detailed characterization. In this context, we investigated gas-phase electrophoresis on a nano electrospray gas-phase electrophoretic mobility molecular analyzer (nES GEMMA, aka nES differential mobility analyzer, nES DMA) as an alternative to standard analytical techniques. In gas-phase electrophoresis, single-charged, surface-dry, native, polydisperse, and aerosolized analytes, e.g., proteins or bio-nanoparticles, are separated according to their electrophoretic mobility diameter, i.e., globular size. Subsequently, monodisperse particles are counted after a nucleation step in a supersaturated atmosphere as they pass a focused laser beam. Hence, particle number concentrations are obtained in accordance with recommendations of the European Commission for nanoparticle characterization (2011/696/EU from October 18th, 2011). Smaller sample constituents (e.g., co-purified proteins) can be detected next to larger ones (e.g., vesicles). Focusing on platelet-derived EVs, we compared different vesicle isolation techniques. In all cases, nanoparticle tracking analysis (NTA) confirmed the presence of vesicles. However, nES GEMMA often revealed a significant co-purification of proteins from the sample matrix, precluding gas-phase electrophoresis of less-diluted samples containing higher vesicle concentrations. Therefore, mainly peaks in the protein size range were detected. Mass spectrometry revealed that these main contaminants belonged to the group of globulins and coagulation-related components. An additional size exclusion chromatography (SEC) step enabled the depletion of co-purified, proteinaceous matrix components, while a label-free quantitative proteomics approach revealed no significant differences in the detected EV core proteome. Hence, the future in-depth analysis of EVs via gas-phase electrophoresis appears feasible. Platelet-derived extracellular vesicles (EVs)with/without additional size exclusion chromatographic (SEC) purification were subjected to nanoparticle tracking analysis (NTA) and gas-phase electrophoresis (nES GEMMA). The latter revealed presence of co-purified proteins, targetable via mass spectrometry (MS). MS also revealed that SEC did not influence EV protein content. To conclude, nES GEMMA is a valuable tool for quality control of EV-containing samples under native conditions allowing for detection of co-purified proteins from complex matrices.

Dataset Information

Analysis of a common cold virus and its subviral particles by gas-phase electrophoretic mobility molecular analysis and native mass spectrometry.

Publications

Analysis of a common cold virus and its subviral particles by gas-phase electrophoretic mobility molecular analysis and native mass spectrometry.

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