Technozym® PAI-1 Antigen ELISA a high sensitivity complete sandwich ELISA for the quantitative determination of Plasminogen Activator Inhibitor type 1 (PAI-1) in human plasma. The assay is a double sandwich ELISA employing two monoclonal antibodies. The system detects active PAI-1, latent and t-PA-PAI complexes. Detection limit is 0.5 ng/ml. Acidified citrated plasma, citrated, CTAD or EDTA plasmas can be used. The assay is standardized with the International standard 87/512. Assay time is 2.5 hours. Kit contents include standards, high and low controls, coated plate, peroxidase-labeled antibody, buffers and ready to use TMB substrate.

  • ELISA test strips: (12), with 8 wells each, coated anti PAI-1 monoclonal antibody; the drying agent is supplied in an aluminium bag.
  • Washing buffer concentrate: (PBS; pH 7.3); containing detergent; 0.01% merthiolate; 1 bottle, 80 ml.
  • Incubation buffer: (PBS; pH 7.3); contains stabiliser protein; 0.05% proclin and dye, 1 bottle, 90 ml, ready for use.
  • Calibrators: (Standards) numbered; lyophilised; 1 bottle each. Concentrations are lot dependent; consult label on the vial.
  • Control plasmas: “low level” and “high level” for checking purposes lyophilised; 1 bottle each. Concentrations are lot-dependent; consult the label on the vial.
  • Conjugate: monoclonal Anti-PAI-1-POX; dyed blue; 1 bottle, 0.3 ml.
  • Chromogen TMB: (tetramethylbenzidine); 1 bottle, 12 ml; ready for use.
  • Stopping solution: sulphuric acid 0.45 mol/l; 1 bottle 12ml; ready for use.
  • Adhesive film: for ELISA test strips (2)
How does a specific gene polymorphism, PAI-1 4G genotype, relate to CVD risk?
For a detailed description of the 4G polymorphism, read Kohler et al. PAI-1 and Coronary Artery Disease. NEJM 2000; 342 (24): 1792-1801. In some studies, the 4G allele (four guanine bases) was significantly associated with high plasma PAI-1 concentrations, and was most strongly associated with previous MI, as well as risk of future MI. Studies have shown that subjects who are homozygous for the 4G allele have plasma PAI-1 concentrations approximately 25% higher than those with the 5G allele (5 guanine bases). Similarly, among patients with hypertriglyceridemia, those with the 4G allele also have higher plasma PAI-1 concentrations than those with the 5G allele. There are still conflicting data on the strength of the relation between PAI-1 gene polymorphism and MI, but it is suggested that the 4G allele is more likely to contribute to MI, particularly in the presence of hypertriglyceridemia.
Won’t tPA be inhibited by PAI-1?
tPA is inhibited in vitro by plasminogen activator inhibitor, so something must be done to avoid this. Acidification of whole blood is therefore performed immediately after withdrawal. This can be done by mixing 1 ml of the citrated blood with 1 ml acetate buffer.
What factors cause increased and decreased levels of t-PA and PAI?
There are numerous physiological factors that influence tPA and PAI antigen and activity level. For a complete list, please request the tPA monograph from DiaPharma. One interesting feature of the fibrinolytic system is the circadian variation in tPA and PAI-1level. Free tPA levels are lowest in the morning, increase during the day, and reach their peak activity level in the late afternoon. tPA and PAI-1 antigen are highest in the early morning and decrease during the day. This may help explain the high incidence of MI and stroke in the morning hours. Other factors that influence tPA and PAI-1 include alcohol, drugs, oral contraceptives, exercise, food, heparin administration, pregnancy, smoking, etc.
What are some of the inhibitors to t-PA?
PAI-1, PAI-2, PAI-3, protease nexin, a2-macroglobulin, trypsin inhibitor, and C1 Inhibitor all inhibit tPA. Plasminogen activator inhibitor 1 (PAI-1) is the most efficient inhibitor of tPA in plasma. It is a serine protease inhibitor (serpin) that acts as a pseudo-substrate for its target protease, with which it forms an inactive complex. PAI-1 is synthesized by several cell types including endothelial cells and hepatocytes and is present in platelets, placenta, and serum. The normal concentration range of PAI-1 in plasma is 5-40 mg/l and the normal activity is 0-20 AU/ml.

PAI-2 is a serpin with a higher affinity for u-PA (urinary-type plasminogen activator, or urokinase) than for t-PA. It is often only detectable during pregnancy, specifically in the third trimester.

PAI-3 is also called Protein C inhibitor, and inhibits u-PA and thrombin, and is present in plasma and urine.

What is the importance of t-PA and PAI in clinical research?
Elevated t-PA and PAI-1 antigen and reduced t-PA activity may be associated with cardiovascular disease. t-PA is given to stroke and heart attack victims shortly after the event to help break up the clot.