Repairing osteochondral defects remains a significant therapeutic challenge due to the complex hierarchical structure of the tissue, while existing treatment strategies are largely palliative rather than curative. For osteochondral tissue production, three-dimensional (3D) bioprinting has become a potent technique to produce biomimetic scaffolds with regulated architecture and cell distribution. This chapter provides a comprehensive and repeatable procedure for the production and biological assessment of 3D bioprinted gelatin methacryloyl (GelMA) scaffolds that incorporate mesenchymal stem cells (MSCs) obtained from bone marrow. The methodology outlines the techniques for photo-crosslinking, extrusion-based bioprinting, MSC expansion and encapsulation, and bioink preparation in order to guarantee cell viability and structural integrity. Comprehensive in vitro evaluation techniques are also described, such as differentiation evaluation related to osteogenic and chondrogenic commitment, cell viability, and proliferation analysis. This methodical approach offers researchers a useful foundation for creating MSC-laden GelMA constructs appropriate for use in osteochondral tissue engineering and additional translational studies.