Tissue-type plasminogen activator (tPA) is a serine protease that activates plasminogen to plasmin in the blood fibrinolytic system. Plasminogen activator inhibitor type 1 (PAI-1) is involved in the regulation of the blood fibrinolytic system and forms a 1:1 covalent complex with tPA and uPA. The sensitive quantitative measurement of total porcine PAI-1 tPA complex antigen in plasma, serum, cell culture media, or tissue extract samples is easily performed with this 96 well strip format ELISA kit. Concentration of PAI-1 tPA complex was found to be 10.2 ng/ml in normal porcine plasma and 37.7 ng/ml in normal porcine serum. The assay measures PAI-1 tPA complex in the 0.2-200 ng/ml range. Free tPA and PAI-1 will not be detected by this assay. Plasma and serum samples should be applied directly to the plate without dilution for best results. Porcine tPA will bind to the capture antibody coated on the microtiter plate. After appropriate washing steps, anti-human PAI-1 primary antibody binds to the captured protein. Excess primary antibody is washed away and bound antibody is reacted with the secondary antibody conjugated to HRP Following an additional washing step, TMB substrate is used for color development at 450 nm. Color development is proportional to the concentration of PAI-1 tPA complex in the samples. A standard calibration curve is prepared in PAI-1 depleted plasma using dilutions of purified PAI-1 tPA complex and is measured along with the test samples. All reagents and standards are provided in these ELISA kits.
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