The purpose of the present study was to assess the effect of magnesium and manganese ions on tissue factor (TF)-induced coagulation and the possible role of factor IX therein. When magnesium chloride or manganese chloride were added in low concentrations to normal human plasma, the human (recombinant) TF-induced coagulation time was shortened. At higher concentrations, magnesium and manganese prolonged the TF-induced coagulation time. Maximum shortening of the coagulation time was obtained at a concentration of 0.5 mmol/l Mn or 2 mmol/l Mg in plasma. Shortening of the TF-induced coagulation time by magnesium and manganese was also observed in factor IX-deficient plasma. A comparison was made between TF preparations from human, rabbit, and bovine brain. The accelerating effect of magnesium was greater with human than with rabbit brain TF. Using bovine brain TF, the clotting time was not shortened by magnesium. Activated factor X-induced coagulation of normal plasma was not accelerated by magnesium. From these experiments, it is inferred that activation of factor X by factor VII–TF can be accelerated by magnesium and manganese ions independently of factor IX.