Abstract: According to the physical parameters such as density, closing flash point and so on, a reference sample with sulfur content of 2.75 wt% was prepared by selecting Merey 16 crude oil as natural matrix. The homogeneity, long-term stability, and storage and transportation stability of the reference samples are tested, and 15 laboratories are selected for determination of sulfur content. The results show that the crude-oil-matrix sulfur reference sample has good homogeneity, long-term stability over 18 months, and storage and transportation stability, which can withstand the temperature of 60 ℃. Its matrix also has good match with samples.