To improve human hepatotoxicity prediction, in vitro liver cell models replicating hepatocyte function, drug metabolism, and toxicity are required. Here, we present a protocol for creating 3D primary human hepatocyte (PHH) cell models using the RASTRUM Platform. We describe the process for PHH model generation; procedures for characterizing the PHH model, including viability, albumin production, and CYP450 inducibility; and drug treatment using acetaminophen and troglitazone. This protocol has applications in upscaling phenotypic hepatotoxicity applications.