Abstract: Objective　To investigate the preparation method of dry blood spots (DBS) control materials for steroids used for internal quality control by liquid chromatography tandem mass spectrometry (LC-MS/MS). Methods　Whole blood was collected and the blood cells and plasma were separated. The blood cells were washed by saline. The activated charcoal was added to the plasma. Standard substance was added to make different concentrations (low, medium, and high) of DBS control materials for steroids. The precision, accuracy, stability, and differences among different blood spots were detected and analyzed by LC-MS/MS. Results　The inter-day precision and accuracy of DBS control materials for steroids were 2.4%-7.0% and 102.0%-111.0%, respectively, and the intra-day precision and accuracy were 5.1%-9.8% and 99.0%-114.8% respectively. The DBS control materials for steroids were stored for 5 months, and there was no difference among the different months (P>0.05). The coefficients of variation among different blood spots were small, 3.3%-8.2%. Conclusions　The DBS control materials for steroids has good precision, accuracy and stability. The difference among different blood spots is small and meet the requirements of indoor quality control products. They can be used for the internal quality control in steroids detection by LC-MS/MS.