The β2-glycoprotein I (β2-GPI) is known for its procoagulant as well as anticoagulant properties. The influence of β2-GPI on tissue factor (TF) activity was investigated by chromogenic assays for both factor Xa generation and factor VIIa activity. When 1.36 mg/mL β2-GPI was added to phospholipids prior to the addition of TF and factor VIIa, the TF activity was moderately inhibited by β2-GPI in a concentration-dependent manner. Inhibition of TF activity by β2-GPI required incubation for at least 20 min. When β2-GPI was added to the mixture of phospholipids and TF followed by the addition of factor VIIa, the TF activity was little affected by the addition of β2-GPI. The TF activity was inhibited when β2-GPI was added after forming a complex with TF and factor VIIa. The results suggest that β2-GPI inhibits both the reconstitution of TF with phospholipids and factor Xa generation by the TF factor-VIIa complex but does not affect the formation of a TF factor-VIIa complex.