High-precision depth-controlled laser bioprinting of cells in extracellular matrix for three-dimensional structures

Bioprinting is an emerging additive manufacturing process that offers great potential for fabricating living tissue by precisely printing cells and biomaterials onto various substrates. This technique can imitate native tissue functions, enabling clinical trials to explore new pathways for regenerative medicine. Among various bioprinting techniques, laser-induced forward transfer (LIFT) offers a high spatial resolution, accurate and controlled bio-ink deposition, and high post-printing cell viability. Effective bioprinting requires a deep understanding of material properties, especially the rheological behavior of bio-inks, which is critical for achieving the desired outcomes. Rheological characterization of these materials is essential for understanding their behavior under bioprinting conditions. The LIFT technique utilizes a wide range of soft biomaterials, generating printed structures containing cells, which proliferate for several days post-printing. These biomaterials can be controllably deposited in a variety of substrates. In this study, two cell-laden bio-inks with low and high number cell densities were printed at controlled depths within an extracellular matrix (ECM) by adjusting the laser energy. This process allows precise immobilization of cells at desired depths within the ECM using light and a proper optical setup. The rheological behavior of all bio-inks was analyzed using a microfabricated rheometer–viscometer on a chip. To investigate the transfer dynamics, a high-speed camera was integrated into the LIFT setup, monitoring the immobilization phenomenon within the ECM, and highlighting important characteristics of the jet propagations during printing. The morphological characteristics of the two sequential and distinct cell-laden jets were examined in detail during the printing process. This study showcases the ability to precisely deposit cells up to 2.5 mm deep within a soft matrix substrate, fabricating any desired cell-laden architecture for bio-engineering applications.