Polydimethylsiloxane (PDMS)-based casting method was used to fabricate PDMS cell culture platforms with molds printed by a fused deposition modeling (FDM) printer. Cell viability study indicated that the produced plates have the suitable biocompatibility, surface properties, and transparency for cell culture purposes. The molds printed from acrylonitrile-butadiene-syrene (ABS) were reusable after curing at 65 °C, but were damaged at 75 °C. To understand thermal damage to the mold at elevated temperatures, the temperature distribution in an ABS mold during the curing process was predicted using a model that considers conduction, convection, and radiation in the oven. The simulated temperature distribution was consistent with the observed mold deformation. As the maximum temperature difference in the mold did not change appreciably with the curing temperature, we consider that the thermal damage is due to the porous structure that increases the thermal expansion coefficient of the printed material. Our study demonstrated that FDM, an affordable and accessible three-dimensional (3D) printer, has great potential for rapid prototyping of custom-designed cell culture devices for biomedical research.