A microplate assay for the screening of ADAMTS-4 inhibitors.

Matrix biology : journal of the International Society for Matrix Biology

PubMedID: 16442274

Thomas M, Sabatini M, Bensaude F, Mignard B, Ortuno JC, Caron I, Boutin JA, Ferry G. A microplate assay for the screening of ADAMTS-4 inhibitors. Matrix Biol. 2006;25(4):261-7.
Aggrecanase plays a major role in cartilage proteoglycan degradation in rheumatic diseases such as osteoarthritis and rheumatoid arthritis. The search of new inhibitors of aggrecanase activity necessitates a robust assays in order to be able to screen large numbers of compounds. We present in this paper an assay based on the cleavage of His-tagged aggrecan interglobular domain by N- and C- terminus truncated, active aggrecanase-1/ADAMTS-4, with formation of the aggrecanase-specific ARGSV neoepitope. This is detected by anti-ARGSV antibody, in turn recognized by a fluorescent anti-IgG. Furthermore, the formation of the reaction products was confirmed by high-pressure capillary electrophoresis. This assay allows the rapid screening of aggrecanase inhibitors in a 96-well plate format, allowing an immediate transposition to high-throughput scale up.