Lidija Filipović - Secondment from ICFC to HPI

Lidija Filipović from partner Innovative Centre Faculty of Chemistry Belgrade spent a month at Hellenic Pasteur Institute, under the EXPAND-EV project.

During the secondment, an antibody-based protocol for extracellular vesicles (EVs) isolation from human plasma using the MojoSort CD81 Microbead Exosomes kit was successfully optimized. Key modifications included replacing the filtration step with low-speed centrifugation (1200 × g, 15 min) to minimize EV loss, and extending bead incubation time from 30 minutes to 1 hour to improve binding efficiency. These changes resulted in a significantly higher EV yield, confirmed by Nanoparticle Tracking Analysis (NTA).Comprehensive hands-on training in NTA was conducted, with a focus on optimizing dilution factors, instrument settings, and data interpretation to achieve accurate, reproducible EV size and concentration measurements. Protein content of isolated EVs was assessed using the Lowry assay; however, due to the inherently low protein levels in EV samples, particle concentration data obtained by NTA were prioritized for downstream proteomic planning.

The secondment provided strong expertise in EV isolation workflow optimization, NTA-based characterization, and preparation of plasma-derived EV samples for proteomic analysis. Close collaboration with consortium partners ensured harmonization of protocols and smooth transfer of samples for proteomics, contributing to high-quality and reproducible clinical EV datasets within WP3.