Antithrombin III is a glycosylated plasma serine protease inhibitor that forms a stoichiometric complex with coagulation cascade enzymes. Antithrombin III inhibits alpha-Thrombin as well as Factor Xa, IXa, XIa and XIIa with heparin enhanced kinetics. Type 1 Antithrombin deficiency is characterized by decreased plasma antigen levels of antithrombin III. The sensitive quantitative measurement of total human antithrombin III antigen in plasma samples is easily performed with this 96 well strip format ELISA kit. An antithrombin concentration of 137 ug/ml in human reference plasma was determined by house testing at 1:100,000 and 1:200,000 dilutions. The assay measures total human antithrombin III in the 0.02-20 ng/ml range. Samples giving human antithrombin III levels above 20 ng/ml should be diluted in blocking buffer before use. A 1:100,000 to 1:400,000 dilution for plasma is suggested for best results. Human antithrombin III will bind to the capture antibody coated on the microtiter plate. After appropriate washing steps, polyclonal anti-human antithrombin III primary antibody binds to the captured protein. Excess primary antibody is washed away and bound antibody, which is proportional to the total antithrombin III present in the samples, reacts with the secondary antibody. Following an additional washing step, TMB substrate is used for color development at 450nm. Color development is proportional to the concentration of antithrombin III in the samples. A standard calibration curve is prepared using dilutions of purified antithrombin III and is measured along with the test samples. All reagents and standards are provided in these ELISA kits.

The Human Antithrombin III standard provided in this kit is calibrated against the WHO International Standard for Human Plasma Antithrombin (NIBSC Code 08/258).