Diapharma PAI-1 Antigen is a two-site ELISA method for the measurement of PAI-1 antigen (Plasminogen-Activator-Inhibitor 1).
Assay designed with two complementary murine monoclonal antibodies specific for PAI-1. The kit contains a calibrator, referenced to the NIBSC international standard, and two control plasmas (low and high). The assay measures homogeneously PAI-1 whether its presentation is: free, active, latent, inactive, complexed with tPA, uPA or vitronectin. Controls are included.
- COAT: Micro ELISA plate, containing 12 strips of 8 wells, coated with a murine monoclonal antibody specific for human PAI-1: Ag, then stabilized; the plate is packed in an aluminium pouch hermetically sealed in presence of a desiccant.
- SD: 2 vials containing 50ml of F-Sample Diluent, ready to use.
- STD: 3 vials of PAI-1 Standard, lyophilized. When restored with 2 ml of F-Sample Diluent, a solution containing about 10 ng/ml of recombinant human PAI-1: Ag is obtained. The exact PAI-IAg concentration is indicated on the flyer provided in the kit.
- CI: 1 vial containing 1 ml of lyophilized Plasma PAI-1 Control I High (human plasma).
- CII: 1 vial containing 1 ml of lyophilized Plasma PAI-1 Control II Low (human plasma).
Note : The PAI-1: Ag concentrations and acceptancy ranges for controls can vary from lot to lot, and are indicated on the flyer provided in the kit.
- IC: 3 vials of Anti-(h)-PAI-1-HRP immunoconjugate, a monoclonal antibody coupled to HRP, lyophilized.
- CD: 1 vial of 25 ml of Conjugate Diluent, ready to use.
- WS: 1 vial of 50 ml of 20 fold concentrated Wash Solution.
- TMB: 1 vial of 25 ml peroxidase substrate: 3,3′,5,5′ – Tetramethylbenzidine containing hydrogen peroxide. Ready to use.
- SA: 1 vial of 6 ml of 0.45M Sulfuric acid (Stop solution). Ready to use.
Note: Use only components from a same kit lot. Do not mix components from different lots, when running the assay.
First, the immunoconjugate, which is a monoclonal antibody specific for PAI-1: Ag coupled to horse radish peroxidase (HRP), is introduced into the microwells coated with another monoclonal antibody specific for PAI-1: Ag. Then, the diluted tested sample is immediately introduced, and the immunological reaction starts. When present, PAI-1: Ag binds onto the monoclonal antibody coated solid phase through one epitope, and fixes the second monoclonal antibody coupled to HRP by another epitope. Following a washing step, the peroxidase substrate, 3,3′,5,5′ – Tetramethylbenzidine (TMB), in presence of hydrogen peroxide (H2O2), is introduced and a blue color develops. When the reaction is stopped with Sulfuric Acid, a yellow color is obtained. The amount of color developed is directly proportional to the concentration of human PAI-1: Ag in the tested sample.