Líffræðifélag Íslands - biologia.is
Líffræðiráðstefnan 2025
Erindi/veggspjald / Talk/poster V32
Höfundar / Authors: Syeda Menahil Fatima (1,2), Jens Guðmundur Hjörleifsson (2), Páll Þór Ingvarsson (1), Erna María Jónsdóttir (1), Annika Heide (3), Berglind Eva Benediktsdóttir(1,4).
Starfsvettvangur / Affiliations: 1. The cancer awareness fund “Göngum saman”, 2. The National Cancer Society Science fund, 3. The Icelandic Student Innovation Fund at Rannís, 4. The University of Iceland Research fund.
Kynnir / Presenter: Syeda Menahil Fatima
Extracellular vesicles (EVs) are cell-derived lipid-bilayer vesicles that transport proteins, lipids, and nucleic acids, regulating essential biological functions. Preserving vesicle size and integrity is crucial for EV isolation methods in therapeutic contexts. Immunoaffinity capture methods utilizing commercially available Fab fragments containing a Twin-StrepTag domain (Fab Streps) directed against EV markers CD9/CD81 has enabled high yield EV recovery with minimal protein contamination and preserved integrity. These Fab fragments bind to StrepTactin affinity resins to enable quasi-immunoaffinity capture. However, these commercial reagents were recently discontinued. This study focused on producing in-house (IH) Fab-Streps to secure supply for academic use. IH CD9 Fab-Streps were overexpressed in E. coli BL21(DE3) and purified using ÄKTA Pure FPLC affinity chromatography yielding high-purity protein after optimizing cultivation temperature/inducer concentration. Effective EV capture with IH CD9 Fab-Strep was confirmed by Western blotting of EV markers CD9 and Alix though particle recovery was 1.5 fold lower than commercial counterparts. For stability assessment, IH CD9 Fab-Streps were freeze-dried and spray-dried with 90% trehalose, remaining aggregation-free and maintaining EV yields comparable to fresh Fab-Streps for at least 60 days under different storage conditions This work ensures Fab-Strep availability for academic research and advances EVs as nanodrug carriers.