Cell culture technology is one of the most core and basic technologies in biotechnology.At present,the limitation and shortcomings of two-dimensional cell culture methods such as multi-well plates,flasks and Petri dishes are obvious.Three-dimensional cell culture can improve the local microenvironment,cell-cell and cell-extracellular matrix interactions,mimic the physiological conditions in vivo to enhance the cell function.Three-dimensional cell culture might be widely used in oncology,drug discovery and other fields in the future.Scaffold is one of the most popular methods for three-dimensional cell culture,which is equivalent to artificial extracellular matrix to mimic the complex three-dimensional structure and the main characteristics of living tissues.This review summarizes three-dimensional cell culture from two aspects:scaffold-based and scaffold-free three-dimensional cell culture,especially introduces the scaffolds.