Multiscale Hybrid Fabrication: Volumetric Printing Meets Two‐Photon Ablation

Abstract

perfusable channels. In the past decades, microfluidic technology has mostly relied on photolithography and soft lithography applied to materials like glass and elastomers (i.e., PDMS), and has been largely limited to 2D devices. [3,6] In recent years, 3D printing has emerged as a powerful tool to generate highly complex, freeform structures with enhanced functionalities, leading to great advances in various fields from photonic crystals to tissue scaffolds. [7][8][9][10][11][12] While two-photon stereolithography has represented the method of choice for the fabrication in the nm to µm scale, [8,9] for constructs with channels in the millimeter to centimeter scale various printing methods have been explored, such as direct ink writing, [13,14] embedded printing with fugitive inks (FRESH, SWIFT), [15][16][17][18] laser-sintering of sacrificial carbohydrate templates, [19] and digital light processing (DLP). [20][21][22] These technologies have particularly contributed to the tremendous progress in the design of increasingly complex vasculature networks for biomedical applications. [18,[23][24][25] More recently, a novel light-based fabrication method termed volumetric printing (VP) has emerged as a promising technology for such applications, enabling the printing of complex centimeter-sized models within seconds. [26,27] Recent studies have demonstrated the possibility to create hollow, perfusable structures, using materials from glass to biopolymers and possibly targeting mesoscale vasculature. [28][29][30][31] However, as all the methods described above, also VP falls short in covering resolution range from µm/sub-µm up to cm, thus currently limiting its application to microfluidic constructs with features >100-200 µm.Another light-based method named two-photon ablation (2PA) instead offers complementary capabilities, being limited in printing time and construct size, but reaching the highest resolution of any biofabrication method (≤1 µm). [8] 2PA is based on multiphoton ionization induced by high-intensity pulsed lasers, [32,33,34] and has been explored for a variety of applications, from "nanosurgery" to form cell-instructive microchannels. [35][36][37][38][39][40][41] Here, we show for the first time the hybrid/combined use of VP and 2PA printing technology to reproduce a multiscale Multiscale printing of 3D perfusable geometries holds great potential for a range of applications, from microfluidic systems to organ-on-a-chip. However, the generation of freeform designs spanning from centimeter to micrometer features represents an unmet challenge for a single fabrication method and thus may require the convergence of two or more modalities. Leveraging the great advances in light-based printing, herein a hybrid strategy is introduced to tackle this challenge. By combining volumetric printing (VP) and high-resolution two-photon ablation (2PA), the possibility to create multiscale models with features ranging from mesoscale (VP) to microscale (2PA) is demonstrated. To successfully combine these two methods, micro...

Embedding biomimetic vascular networks via coaxial sacrificial writing into functional tissue

“…1 Without the ability to embed immediately addressable and perfusable vasculature, these engineered human tissues do not remain viable over the time required to provide therapeutic benefit. [2–7] Recent advances in extrusion [8–10] , embedded [3,11–14] , and light-based [15–17] bioprinting have begun to address this critical need. Yet no method currently allows the free-form patterning of hierarchical, branching vasculature composed of smooth muscle cell-laden shells that surrounds endothelialized lumens in acellular or densely cellular tissue matrices.…”

Virtual Volumetric Additive Manufacturing (VirtualVAM)

“…This approach overcomes important limitations of traditional AM methods, where layer‐by‐layer addition of material hinders production rates, part quality, and geometric flexibility. More broadly, VAM is capable of rapid production of complex parts in a broad range of polymeric materials (e.g., high viscosity acrylates, [ 1 ] thiol‐enes, [ 4,5 ] cell‐laden hydrogels, [ 6,7 ] preceramic, [ 8 ] and glass‐forming [ 9 ] polymer resins).…”

Crafting Tissue Complexity

“…Rizzo et al, demonstrated the feasibility of combining the high-speed printing of large VBP constructs with perfusable macrochannels with the subsequent 2-photon ablation of high-resolution (∼2 mm) microchannels. [614] The resulting convoluted and multi-scale tubular networks greatly resemble native vascular structures, but given the short working depth of the 2-photon ablation process (∼500 mm), such a converged approach will be challenging to upscale. In its current form, such an approach can only be used to pattern thin constructs of the border regions of larger VBP-produced structures.…”

“…In studies demonstrating this approach, the 3D resolution of grafted structures is significantly enhanced, given the design freedom in across all planes that is possible in this approach. However, as with several multi-photon approaches, the working depth of this approach is rather limited (< 1 mm for various studies employing hydrogels) [614,615,643] and approximately 2-3 mm for microscopy applications. [644,645] This means that convoluted 3D printed structures of clinically relevant sizes could not be biofunctionalized with full spatial freedom.…”

“…Volumetric printing of cellular structures (volumetric bioprinting), has been demonstrated on various occasions, reporting the possibility to print viable and functional cells and organoids, [127,286] and to sculpt the latter into complex, function-modulating architectures. [286] The potential to create complex multi-material Thermal Shrinking of Biopolymeric Hydrogels for High Resolution Volumetric Printing A constructs, [473] combine the VP approach with other printing strategies, like embedded bioprinting for further multi-material and multi-cellular applications, [474] two-photon ablation to create microchannels withing VP-printed structures, [614] and melt electrowriting to enhance the mechanical stability of printed hydrogel structures, [472] has also been recently demonstrated. In the field of material chemistry however, the use of smart, stimuli responsive bioresins has been limited to date.…”