Horses, like humans, can experience bone fractures and due to their large size and need to bear weight on all limbs during the recovery period, they can be difficult to treat. Surgical techniques to improve fracture repair are improving, but to date, regenerative medicine technologies to aid fracture healing are not commonly applied in horses. We have previously demonstrated that equine induced pluripotent stem cells (iPSCs) can be differentiated into bone forming osteoblasts in 2D culture. Here we report on the use of a thermoplastic, 3D printed polymers to provide a scaffold for successful, in vitro osteoblast differentiation of equine iPSCs. The scaffolds provides a transparent, cost effect solution to allow the analysis of osteoblast differentiation using live cell imaging, immunohistochemistry and quantitative PCR. This in vitro study demonstrates the future feasibility of generating 3D bone constructs through the cell seeding of scaffolds to use in regenerative medicine strategies to work demonstrates the possibility of using cell-based therapies in combination with scaffold technologies for improving improve fracture repair in a relevant, large animal model.