Abstract: In order to obtain a higher content of Foot-and-mouth disease virus（FMDV） under the low-serum whole suspension culture of BHK-21 cells, the culture characteristics of 4 strains of BHK-21 suspension cells from different sources were compared. O-type and A-type FMDV were inoculated at 1%, 2% and 5% of the volume, respectively, and samples were taken at 4, 8, 12, 16, 20, and 24 h after inoculation for comparative study of 146 S and median lethal dose(LD₅₀). BHK-21 suspension cell line with good morphology and more stable culture characteristics was screened; BHK-21 suspension cell line was screened by shake flask suspension culture. Using O-type FMDV O/HB/HK/99 strain and A-type FMDV AF/72 strain, the study was carried out according to the effect of inoculation dose, viable cell density（VCD）, virus culture temperature, virus culture pH value, medium exchange ratio and other process parameters on virus 146 S expression. And in the 7.5,50,500 L bioreactors, it was amplified step by step, and 3 batches of virus suspension culture of O-type FMDV O/HB/HK/99 strain and A-type FMDV AF/72 strain were carried out; the culture process of O-FMDV O/HB/HK/99 strain and A-FMDV AF/72 strain inoculated with BHK-21 suspension cells was determined. The results showed that a BHK-21-CP005 suspension cell line with good morphology and more stable culture characteristics was screened out. The optimal conditions for the proliferation of O-type FMDV O/HB/HK/99 strain and A-type FMDV AF/72 strain in BHK-21-CP005 suspension cell line cultured in low serum suspension in shake flasks were as follows: when the cells were cultured for 48 h, the VCD reached 4.5 ×10~6/mL; O/HB/HK/99 strain and AF/72 strain were inoculated at 2% by volume; the culture temperature was 36.5-37.0 ℃; the pH value was 7.30-7.50; the medium was changed by 4∶5 in volume; the virus solution was harvested 16-20 hours after virus inoculation; the content of virus 146 S was not lower than 9.0 μg/mL, and the LD₅₀ was not lower than 1×10^8.50/0.2 mL. The results suggested that the use of BHK-21-CP005 strain low serum suspension culture to produce higher content of FMDV had a stable process and could realize large-scale production.