AssayPro—Human Thrombin ELISA

Introduction
Thrombin (activated Factor II [IIa]) is a coagulation protein that has many effects in the coagulation cascade. Thrombin is a serine protease (EC 3.4.21.5) that converts soluble fibrinogen into insoluble strands of fibrin, as well as catalyzing many other coagulation-related reactions (1). Thrombin is in the form of alpha-thrombin that is the immediate end product of prothrombin activation: two further thrombin products can be identified, beta- and gamma-thrombin. These are degraded forms that may arise from autodigestion of a thrombin preparation (2, 3).

Principle of the Assay
The AssayMax Human Thrombin ELISA kit is designed for detection of alpha- thrombin in human cell culture supernatants. This assay employs a quantitative sandwich enzyme immunoassay technique that measures thrombin in less than 4 hours. A monoclonal antibody specific for thrombin has been pre-coated onto a microplate. Thrombin in standards and samples is sandwiched by the immobilized antibody and a biotinylated polyclonal antibody specific for thrombin, which is recognized by a streptavidin-peroxidase conjugate. All unbound material is then washed away and a peroxidase enzyme substrate is added. The color development is stopped and the intensity of the color is measured.