Protocol for generating an in vitro 3D multicellular culture model of ovarian high-grade serous carcinoma

The development of translational ovarian cancer models to investigate and overcome treatment resistance, while accounting for the impact of the tumor microenvironment, is critical. Here, we present a protocol to establish a multicellular culture model that retains both genetic complexity and the microenvironment of patient tumors and is amenable to molecular and phenotypic analyses and high-throughput drug testing. We describe steps for culturing and characterizing stromal cells derived from cryopreserved and fresh samples and detail procedures for combining them with organoids.