Capillary zone electrophoresis of bacterial extracellular vesicles: A proof of concept
Martyna Piotrowska, Krzesimir Ciura, Michalina Zalewska, Marta Dawid, Bruna B. Correia, Paulina Sawicka, Bogdan Lewczuk, Joanna Kasprzyk, Laura Solá, Wojciech Piekoszewski, Bartosz Wielgomas, Krzysztof Waleron, Szymon Dziomba
Abstract
The extracellular vesicles (EVs) released by plant pathogens of the Pectobacterium genus were investigated. The isolates were obtained using differential centrifugation followed by filtration and were characterized in terms of total protein content and particle size distribution. The transmission electron microscopy (TEM) analysis revealed the presence of two morphologically differentiated subpopulations of vesicles in the obtained isolates. The proteomic analysis using matrix-assisted laser desorption ionization mass spectrometry with time of flight detector (MALDI-TOF/TOF-MS) enabled to identify 62 proteomic markers commonly found in EVs of Gram-negative rods from the Enterobacteriaceae family. Capillary electrophoresis (CE) was proposed as a novel tool for the characterization of EVs. The method allowed for automated and fast (<15 min per sample) separation of vesicles from macromolecular aggregates with low sample consumption (about 10 nL per analysis). The approach required simple background electrolyte (BGE) composed of 50 mM BTP and 75 mM glycine (pH 9.5) and standard UV detection. The report presents a new opportunity for quality control of samples containing EVs.