Gas-phase electrophoresis of exhaled breath condensate (EBC) applying a nES GEMMA instrumentation

Abstract

Gas-phase electrophoresis of exhaled breath condensate (EBC) applying a nES GEMMA instrumentation

Victor U. Weiss

TU Wien, Institute of Chemical Technologies and Analytics, Vienna, Austria Email: victor.weiss@tuwien.ac.at

Introduction Exhaled breath condensate (EBC) is a promising, non-invasive, diagnostic sample obtained by condensation of exhaled breath. In contrast to nasal or throat swabs, bronchoalveolar lavage, sputum or saliva, EBC is easily accessible in large quantities. Hence, it can be applied to target the protein and (bio-)nanoparticle content released from the respiratory tract of an individual. It was the aim of our research to characterize EBC in this respect by application of gas-phase electrophoresis on a nano Electrospray Gas-phase Electrophoretic Mobility Molecular Analyzer (nES GEMMA) setup.

Methods Gas-phase electrophoresis on a nES GEMMA also known as nES Differential Mobility Analyzer (nES DMA) separates single-charged, aerosolized, surface-dry analytes in the gas-phase at ambient pressure according to the particle size [1]. Previous experiments have demonstrated the applicability of such a setup for the characterization of proteins and protein aggregates, viruses, virus-like particles, liposomes, extracellular vesicles and similar (bio-)nanoparticles. Besides information on surface-dry particle size (EM diameter), analyte and sample heterogeneity and number-based particle concentrations, information on particles’ molecular weight values can be gathered by application of a respective correlation [2].

Results EBC was collected from a voluntary donor. Subsequently, protein and (bio-) nanoparticle content of EBC was enriched by evaporation of water in vacuum. Residual components were taken up in 40 mM ammonium acetate, pH 8.4 as volatile electrolyte, necessary for the nES process preceeding gas-phase electrophoresis. nES GEMMA of corresponding samples demonstrated high amounts of non-volatile, low molecular weight components in samples (salts, etc.). Removal of these components was assessed via spin filtration (10 kDa MWCO filters, polyethersulfone membrane) and dialysis (10 kDa MWCO, modified cellulose membrane). Only for the latter case, nES GEMMA showed signals for proteins and (bio-)nanoparticles. Upon spin filtration, these components were lost probably due to interaction of analytes with the membrane. UV Vis absorption and nanoparticle tracking analysis were applied to follow individual sample preparation steps. To conclude, the presented work demonstrates the combination of EBC collection and gas-phase electrophoresis on a nES GEMMA instrument to characterize the protein and (bio-)nanoparticle content obtained from the respiratory tract of an individual in a proof of concept study.

Innovative aspects • EBC collection and subsequent sample preparation for nES GEMMA is demonstrated • UV Vis and NTA measurements were applied to follow individual sample preparation steps • nES GEMMA reveals protein and nanoparticle content in EBC in the low nm range

References [1] Kaufman, S.L., Skogen, J.W., Dorman, F.D., Zarrin, F. and Lewis, K.C. (1996) Anal. Chem., 68 (11), 1895–1904. [2] Bacher, G., Szymanski, W.W., Kaufman, S.L., Zöllner, P., Blaas, D., and Allmaier, G. (2001) J. Mass Spectrom., 36 (9), 1038-1052.