We immunohistochemically examined the expression of matrix metalloproteinase (MMP)-2 and MMP-9 using venous thrombi developed by ligation of the inferior vena cava (IVC) in mice. Both MMP-2- and MMP-9- positive cells could be detected in the whole course of thrombus formation after IVC ligation. Morphometrically, their number was greatest 14 days after IVC ligation and thereafter, gradually decreased at 21 days. The number of MMP-9-positive cells was significantly higher than that of MMP-2-positive cells at 1 to 7 days. The average ratio of MMP-9 to MMP-2 (MMP-9/MMP-2 ratio) was >2.0 in all thrombus samples at 1-5 days. After 7 days, the MMP-9/MMP-2 ratio was less than 2.0. These observations implied that an MMP-9/MMP-2 ratio markedly exceeding 2.0 strongly indicates an age of 5 days or less. Furthermore, an MMP-9/MMP-2 ratio of <2.0 probably indicates an age of more than 7 days. The present study demonstrated that the immunohistochemical detection of intrathrombotic MMP-2 and MMP-9 was suitable to estimate the age of venous thrombi.