Use of EIA to Measure MMPs and TIMPs

The activity of matrix metalloproteinases (MMPs) is closely regulated by tissue inhibitors of metalloproteinases (TIMPs) under pathophysiological conditions. The quantitative imbalance between MMPs and TIMPs in tissues and body fluids is considered to cause tumor invasion, metastasis, and various connective tissue diseases such as arthritis and periodontitis (1 ). MMPs and TIMPs have been identified and purified from various samples such as cell-conditioned media, tissue homogenates, sera, and so on. Furthermore, monoclonal antibodies against MMPs and TIMPs have been also prepared, and enzyme immunoassays (EIAs), which can quantitate MMPs and TIMPs in specimens, have been established. A two-step sandwich EIA for human MMP-3 was prepared using the combination of a monoclonal antibody as the solid-phase with rabbit polyclonal antibody and horseradish peroxidase (HRP)-labeled donkey antirabbit gamma globulin as the second phase (2 ). EIAs for human MMP-9 were developed using MMP-9 specific polyclonal antibody (3 ,4 ) or monoclonal antibodies (5 ). For quantitative determination of MMP-8, a two-step sandwich EIA was developed using polyclonal antibody (6 ). We also established one-step sandwich EIAs for MMPs and TIMPs using two kinds of monoclonal antibodies (7 –16 ).