For decades, scientists have attempted in vitro culture of marine sponge cells and met with little success. Recently, the use of an optimized nutrient medium in two-dimensional cell culture has produced rapid cell division in three species of sponges in the genus Geodia. Continued division after the cells have formed a confluent monolayer has not yet been achieved. In addition, evidence shows that cells that are cultured in a two-dimensional environment exhibit changes in morphology and functionality. The use of three-dimensional cell culture techniques may increase the number of cells cultured in vitro by providing more surface area for the cells to adhere to. It may also promote normal functioning of the cells through increased cell-to-cell and cell-to-extracellular matrix communication, which are believed to perform a key role in cell functionality and morphology. This research combines the use of optimized medium with three-dimensional cell culture methods to culture cells of the marine sponge Geodia neptuni. Optimization of the culture methods demonstrated here may lead to the ability to scale up in vitro culture of biomedically important marine sponge species for the production of marine natural products.