Development of an automated method for 3D Tissue Engineering

The Federal Drug Administration (FDA) recently eliminated the mandatory use of animal testing in the drug approval process, a decision that coincides with similar regulatory changes in Europe. In line with these changes, the European Parliament has actively supported research projects aimed at reducing animal testing. These initiatives are driven by societal demand for more ethical research practices and are part of a broader movement toward ethical scientific research. In this evolving landscape, in vitro test systems are emerging as valuable tools. These cell-based models adhere to the 3Rs principles – Replace, Reduce, and Refine – and show significant potential to replace traditional animal testing methods. This study presents "Robotic enabled biological Automation - ReBiA", an innovative automated system that transforms manual laboratory processes into automated equivalents through the use of a dual-arm robot. As part of this work, all the tools and subsystems required for the culture of 3D in vitro models were identified. While automated versions of subsystems like the centrifuge, single-channel pipettes and microscope could be purchased and integrated directly into the central control system, dedicated liquid storage systems and serological pipette tips were designed. In addition, a serological pipette was developed that can be used by the robot to effectively transfer volumes greater than 5 ml and remove media supernatants. After technical installation, the system was successfully subjected to a hygiene test. Subsequently, the ReBiA system has demonstrated its efficacy by successfully producing complex three-dimensional in vitro test models, including reconstructed human epidermis (RHE) and human airway models. The application of the automated RHE was tested in an irritation test according to the Organization for Economic Cooperation and Development (OECD) guideline 439. In addition to the fabrication of membrane-based tissue models, the ReBiA system demonstrated its suitability for the automated culture and analysis of tumor spheroid therapy studies. By using experimental designs to test different combinations of alginate-collagen hydrogels, the study also shows that automated processing of hydrogels of different viscosities is possible with the hydrogel dosing unit. This proves that ReBiA can support the systematic development of new materials for Tissue Engineering. In addition, the handling of the temperature-sensitive hydrogel Matrigel® was implemented. Using intestinal organoid cultures as an example, a proof-of-concept study showed that ReBiA is able to deliver cell-laden Matrigel® in a defined manner and to manage the liquid media for this type of culture. ReBiA fuses computer science, engineering, and biology to create a sophisticated platform for rapid implementation of new Tissue Engineering processes in a compact 2.5 x 2.5 m footprint. The variety of protocols successfully realized with this system underscores its flexibility and confirms the hypothesis that a centrally automated solution with a dual-arm robot is exceptionally well suited to provide a versatile platform for the field of Tissue Engineering.