Contents of Kit
1. Capture Antibody (yellow): 0.5 ml of polyclonal affinity purified anti-tPA antibody for coating plates.
2. Detecting Antibody (red): 0.5 ml of peroxidase conjugated polyclonal anti-tPA antibody for detection of captured tPA.
Note: Reagents are sufficient for at least 5×96 well plates using recommended protocols. Antibodies are supplied in a 50% (v/v) glycerol solution for storage at –10 to -20°C. Keep vials tightly capped. Do not store in frost-free freezers.
Tissue-type plasminogen activator (tPA) is one of two major physiologic activators of plasminogen in plasma. It is a serine protease of 68 kDa produced primarily in endothelial cells but is also present in monocytes and megakaryocytes. Normal plasma tPA antigen concentrations have been reported from 20 ng/ml to 5 μg/ml, depending on the assay used, but typically most of the tPA (> 90%) is in complex with it's primary inhibitor, plasminogen activator inhibitor-1 (PAI-1). Structurally, tPA is a single-chain enzyme that consists of a catalytic domain followed by two kringle structures, an EGF domain and a finger domain. The activation of plasminogen by tPA is dependent on the presence of a fibrin cofactor. The binding of both tPA and plasminogen to fibrin is mediated in part through lysine binding sites within the kringle structures of both enzyme and substrate, but also through the finger domain of tPA. Activation of plasminogen by tPA occurs by cleavage after residue Arg560 to produce the two-chain active serine protease plasmin. The activity of tPA is regulated in part by a very short half life in circulation (t½ of ~4 minutes) and by circulating protease inhibitors PAI-1 and to a lesser extent α2 macroglobulin.