Scalable and Controlled Production of Microalgae-derived Extracellular Vesicles Using a Differential Tangential Flow Filtration Protocol.

The transition of extracellular vesicle (EV) production from laboratory to industrial scale presents major challenges in terms of standardization, scalability, and product consistency. Here, we present a scalable manufacturing strategy for nanoalgosomes, EVs derived from the microalga Tetraselmis chuii. The process integrates upstream cultivation under controlled conditions with downstream isolation and purification via continuous tangential flow filtration (TFF). Critical TFF parameters-cross-flow velocity, shear rate, permeate flux, and transmembrane pressure-were investigated to evaluatetheir impact on process behaviour, extracellular vesicle recovery, and functional integrity. Following the MISEV2018 guidelines established in our previous baseline characterizations, the isolated nanoalgosomes were verified by a quality control, including biochemical and biophysical characterization, confirming the robustness of the platform. Overall, this work provides a practical processing framework for the scientific community dealing with large-volume EV bioprocessing, offering an informative baseline for the scalable production of microalgae-derived extracellular vesicles.