Srishti: a custom-built laser-assisted 3D bioprinter for fabricating high-resolution biological constructs

Abstract

Laser-assisted bioprinting is a nozzle-free, non-contact technique that employs drop-on-demand technology for the fabrication of high-resolution three dimensional (3D) structures for various applications including tissue engineering, in vitro modeling, spheroid/organoid production, biosensors, and more. Globally, custom-designed laser bioprinters are primarily being developed to create tissue-engineered cellular constructs. Bioink printability and its printing resolution are influenced by the design of the laser bioprinter and its parameters. There are only a few established laser bioprinters that are actively employed in the to development of living cellular constructs. “Srishti”, a custom-built laser-assisted bioprinting system with 8-axes control movement, was specifically designed, and developed to explore diverse biomedical applications. It is designed to facilitate precise control over motor movements to achieve high levels of reproducibility. This study examined the influence of key parameters such as donor-receiver distance (mm), donor & receiver printing speed (mm/s), bioink concentration (% & w/v), and bioink coating thickness (µm) on laser bioprinting of pre-designed patterns. Laser parameter optimization resulted in a significant enhancement of printed line width resolution, reducing it 13-fold from 1000 ± 47 μm to 78 ± 8 μm. Various bioink formulations, including sodium alginate, gelatin, and sodium alginate/gelatin blends, were investigated to determine the ideal bioink properties required for laser bioprinting. Further, in vitro studies were carried out to determine the viability of NCTC clone 929 (mouse fibroblast) cells after laser bioprinting. Under optimized conditions, bioink droplets containing one to three cells with an average diameter of 58 ± 6 μm were successfully printed using the Srishti bioprinter. Additionally, embryonic rat cardiomyoblast (H9c2) cells were also bioprinted to demonstrate viability upto 8 days in culture. Our analysis reveals the successful development of a cost-effective laser bioprinting system that offers precise control and facilitate printing of high resolution cellular bioink droplets. The designed Srishti system has been proven to be effective in live cell patterning with excellent viability and expression native cell phenotypes post-printing. These findings highlight the potential of Srishti in fabricating high-resolution tissue structures, making it a promising system for applications in tissue engineering, and drug testing.