Two-dimensional (2D) cell culture systems are normally used for in vitro experiments. These systems have some drawbacks which affect results of in vitro experiments because cultured cells have different morphology and physiological properties comparing to in vivo models. Therefore, there are current technique to develop three-dimensional (3D) cell culture systems which are closer to the environment in real tissues. In our study, we used polycaprolactone fiber scaffold for culture of bovine monocytes to develop dendritic cells (DCs) from those cells. Cultured monocytes in the polycaprolactone nanofiber scaffold differentiated to DCs. The differentiated DCs showed a 3D structure in the scaffold. These results suggest that this method is suitable for the development of DCs in a manner of three-dimension. These methods are perspective for bovine cell culture and has advantages compare to conventional 2D culture systems.