Fabricating heterogeneous alginate microbead arrays with spatially prescribed distributions using laser direct-write bioprinting

Abstract

Manufacturing 3D tumor systems with control over the spatial placement of disparate cell types (e.g., tumor, stromal) has posed a significant challenge. Compared to other fabrication methods, bioprinting approaches are particularly well suited to the task because they offer the fabrication of high-resolution constructs with high spatial control. Inkjet bioprinting represents the benchmark bioprinting approach because it can print multiple live cells and rapidly generate intricate patterns with high-throughput fabrication. However, inkjet bioprinting has limited resolution, increased potential for clogging cell-loaded bioinks, and could potentially introduce high shear stresses to cells. Alternatively, LDW bioprinting enables nozzle-free, noncontact fabrication of cells and cell-loaded microbeads with high resolution and spatial control. Further, LDW can create and pattern size-controlled 3D cell microenvironments (i.e., microbeads, microcapsules) in a single step. Herein, this work addresses the fabrication of spatially heterogeneous microbead arrays across multiple bioinks via LDW bioprinting. Two methods for bioprinting multiple bioinks onto the same substrate were explored, utilizing untagged and fluorescent-tagged alginates, demonstrating potential approaches to fabricate spatially heterogeneous constructs. LDW showed accurate and precise microbead placement for creating constructs with prescribed spatial composition. Further, spatially heterogeneous constructs were fabricated with individual microbead placement and guided by either user-defined idealized geometries or histologic image templates. This platform provides critical foundations for fabricating heterogeneous 3D tumor models that better reflect patient heterogeneities.