This experiment was carried out to determine the progesterone concentration of bovine plasma by liquid phase double antibody enzyme immunoassay. The optimum conditions of assay-system, double (first and second) antibody and carrier (normal rabbit serum) were investigated. The optimum dilution rate of first antibody, second antibody and normal rabbit serum was $10{\times}10^3$ to $15{\times}10^3$, 20 and $1{\times}10^3$ times, respectively.