Project Summary The purpose of this proposal is to advance the field of Mesenchymal Stromal Cell (MSC) biology and particularly the process of translating MSCs into safe and effective therapies. The Broad Aim of this proposal is to demonstrate that the Cell X™ automation platform enables users to generate culture expanded human “mesenchymal stem/stromal cell” populations (hMSCs) with greater quality and with less variation from batch-to-batch. Success will open the valued market of MSC generation and commercial fabrication to Cell X™ Technologies. The Cell X™ Device is the core product of Cell X Technologies, which was founded in 2017 as a spin-off company derived from technology generated in the Muschler Lab at the Cleveland Clinic with engineering support from Parker Hannifin Corporation. Cell X combines four essential automated features: 1) Rapid large field of view (LFOV) (“Google Earth” whole well) image analysis, enabling the user to quantitatively capture and use “performance-based” colony (clone) attributes for cell source selection. 2) Precise automated methods for biopsy, picking or weeding (removal) of colonies with specific attributes. 3) Automated methods for removal and replacement of media during cell expansion (i.e. media change). 4) Consistent and standardized documentation of parameters and images associated with each intervention. hMSC variation represents a profound point of ongoing “pain point” in the biomedical research community and in the emerging industry of regenerative medicine and cellular therapies. Clinical use demands high levels of repeatability and reproducibility. Donor-to-donor and batch-to-batch variation also confounds the utility of culture expanded MSCs in drug screening, toxicity testing, evaluation of biomaterials, etc. Demonstration of the value of Cell X™ automation for optimizing MSC generation and fabrication will greatly advance the value and demand for Cell X™ Platforms for research, drug development and clinical fabrication.