Abstract: Tumor is a complex system of tumor cells, fibroblasts, tumor blood vessels, immune cells, extracellular matrix and other components. In recent years, multicellular tumor spheroid is increasingly used as 3D in vitro tumour model. Compared to 2D culture model in vitro, it can better character fundamental structure and microenvironment of tumour tissue in vivo. It is playing an important role in antitumour drug screening test and tumour biology. In this paper, we make a review of the development of multicellular tumour spheroid and its application in drug screening and preparation evaluation.