Characterizing the Process Physics of Ultrasound-Assisted Bioprinting.

3D bioprinting has been evolving as an important strategy for the fabrication of engineered tissues for clinical, diagnostic, and research applications. A major advantage of bioprinting is the ability to recapitulate the patient-specific tissue macro-architecture using cellular bioinks. The effectiveness of bioprinting can be significantly enhanced by incorporating the ability to preferentially organize cellular constituents within 3D constructs to mimic the intrinsic micro-architectural characteristics of native tissues. Accordingly, this work focuses on a new non-contact and label-free approach called ultrasound-assisted bioprinting (UAB) that utilizes acoustophoresis principle to align cells within bioprinted constructs. We describe the underlying process physics and develop and validate computational models to determine the effects of ultrasound process parameters (excitation mode, excitation time, frequency, voltage amplitude) on the relevant temperature, pressure distribution, and alignment time characteristics. Using knowledge from the computational models, we experimentally investigate the effect of selected process parameters (frequency, voltage amplitude) on the critical quality attributes (cellular strand width, inter-strand spacing, and viability) of MG63 cells in alginate as a model bioink system. Finally, we demonstrate the UAB of bilayered constructs with parallel (0°-0°) and orthogonal (0°-90°) cellular alignment across layers. Results of this work highlight the key interplay between the UAB process design and characteristics of aligned cellular constructs, and represent an important next step in our ability to create biomimetic engineered tissues.