3D-printed ordered bed structures for chromatographic purification of enveloped and non-enveloped viral particles

Abstract In the last few decades we have witnessed a rapid advance in the design of new biotherapeutic products using viral particles. The current limitations of manufacturing these particles are often associated with their purification processes. Additive manufacturing opened the possibility of printing 3D porous chromatographic structures to be used in the bio-separation field. Here, we report the production and use of 3D-printed cellulose chromatographic columns functionalized with two different ligands: diethylaminoethyl (DEAE) and hydroxyapatite for the purification of oncolytic adenovirus and lentiviral vectors. Porous beds with ordered channel structures can be manufactured to achieve separations that are efficient and tailored to the target products. While common chromatographic beads are based on non-ordered bed structures, in this work we explore the use of a triply periodic minimal surface, the Schoen Gyroid, to create bed structures with a channel size of 300 µm in diameter. The dynamic binding capacity (DBC) obtained for non-enveloped oncolytic adenovirus using the 3D-printed DEAE column (1.9 × 1010 viral genomes per millilitre) was consistent with the reported values in the literature for this virus particle. We also demonstrate that by using these 3D-printed chromatographic supports, oncolytic adenoviruses were successfully purified with a recovery yield of 69 ± 6%, while maintaining their size and shape as observed by electron microscopy analysis. For lentiviral vectors, a DBC of 2 × 109 physical particles per millilitre was achieved for the DEAE column. Moreover, these enveloped viruses were purified using this 3D-printed column with a recovery yield of 57% of transducing units. These findings suggest, with advancements, 3D printing technologies applied in the viral vector manufacturing field could improve downstream process efficiency.