Optimized Tangential Flow Filtration for Efficient Isolation of Highly Pure and Bioactive sEVs.

Area of Science:

• Biotechnology
• Nanomedicine
• Cell Biology

Background:

• Small extracellular vesicles (sEVs) hold significant therapeutic promise but scalable isolation methods are lacking.
• Tangential flow filtration (TFF) is a potential technique for large-scale sEV separation, yet optimal parameters require systematic investigation.

Purpose of the Study:

• To investigate the impact of key TFF parameters on sEV separation and purity.
• To identify optimal TFF conditions for efficient and scalable sEV isolation.

Main Methods:

• Systematic evaluation of TFF parameters: shear stress, MWCO, TMP, and washing cycles.
• Characterization of sEVs using particle concentration, size, morphology, and protein analysis.
• Comparison of TFF-isolated sEVs with ultracentrifugation-isolated sEVs.

Main Results:

• Excessive shear stress (>9.05 Pa) caused sEV damage and protein aggregation.
• A 750 kDa MWCO membrane and 3-5 psi TMP minimized sEV leakage and maximized filtration.
• Optimized TFF yielded sEVs with comparable purity to ultracentrifugation but with higher recovery and bioactivity.

Conclusions:

• Optimized TFF parameters enable efficient sEV purification, balancing purity, recovery, and bioactivity.
• This optimized method facilitates scalable sEV production for therapeutic applications.
• The findings accelerate the clinical translation of sEV-based therapeutics.