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Subunit vaccines stand as a leading approach to expanding the current portfolio of vaccines to fight against COVID-19, seeking not only to lower costs but to achieve long-term immunity against variants of concern and have the main attributes that could overcome the limitations of the current vaccines. Herein a chimeric protein targeting S1 and S2 epitopes, called LTp50, was designed as a convenient approach to induce humoral responses against SARS-CoV-2. LTp50 was produced in recombinant Escherichia coli using a conventional pET vector, recovering the expected antigen in the insoluble fraction. LTp50 was purified by chromatography (purity > 90%). The solubilization and refolding stages helped to obtain a stable protein amenable for vaccine formulation. LTp50 was adsorbed onto alum, resulting in a stable formulation whose immunogenic properties were assessed in BALB/c mice. Significant humoral responses against the S protein (BA.5 variant) were detected in mice subjected to three subcutaneous doses (10 µg) of the LTp50/alum formulation. This study opens the path for the vaccine formulation optimization using additional adjuvants to advance in the development of a highly effective anti-COVID-19 vaccine directed against the antigenic regions of the S protein, which are less prone to mutations.

Two-photon stereolithography (TPS) is an established additive fabrication technique allowing the voxel-by-voxel direct writing of even intricate 3D nano/microstructures via the polymerization of a photoresin. An obvious way to tune the chemical functionalities of such nano/microstructures is formulating a photoresin with the desired functional monomer(s). Unfortunately, this makes every photoresin "unique" in terms of viscosity and reactivity, thus requiring a tedious and often time-consuming optimization of its printing parameters. In this work, we describe a general approach for the chemical functionalization of TPS-written structures based on two commercial photoresins. Our strategy entailed the grafting of functional polymer layers via an innovative approach based on photoiniferter coupling to unreacted double bonds and photopolymerization. After writing woodpiles as 3D model structures, we demonstrated the viability of this approach by anchoring a photoiniferter via its photoinduced addition to the residual CC on the structure's surface triggered by green light. This in turn allowed for the blue light-mediated, surface-initiated photopolymerization of functional monomers. Molecularly imprinted polymer films were also easily synthesized by using the same approach on model honeycombs. The imprinted layers resulted in only a minimal increase in size with no effect on the geometrical features of the honeycombs. Overall, this strategy offers a general approach for the surface modification of TPS-written (meth)acrylic structures with a wide variety of functional polymers via photoiniferter polymerization.