Description

Nori Mouse Serpin E1/PAI-1 ELISA Kit Summary

Alternative names for serpin E1: Plasminogen activator inhibitor-1 (PAI-1), endothelial plasminogen activator inhibitor, serpine1, Mr1, Pai1, Planh1

Standard Curve

Kit Components
1. Pre-coated 96-well Microplate
2. Biotinylated Detection Antibody
3. Streptavidin-HRP Conjugate
4. Lyophilized Standards
5. TMB One-Step Substrate
6. Stop Solution
7. 20 x PBS
8. Assay Buffer

Other Materials Required but not Provided:
1. Microplate Reader capable of measuring absorption at 450 nm
2. Log-log graph paper or computer and software for ELISA data analysis
3. Precision pipettes (1-1000 µl)
4. Multi-channel pipettes (300 µl)
5. Distilled or deionized water

Protocol Outline
1. Prepare all reagents, samples and standards as instructed in the datasheet.
2. Add 100 µl of Standard or samples to each well and incubate 1 h at RT.
3. Add 100 µl of Working Detection Antibody to each well and incubate 1 h at RT.
4. Add 100 µl of Working Streptavidin-HRP to each well and incubate 20 min at RT.
5. Add 100 µl of Substrate to each well and incubate 5-30 min at RT.
6. Add 50 µl of Stop Solution to each well and read at 450 nm immediately.

Background: 

Serpin E1 also named Plasminogen activator inhibitor-1 (PAI-1) or endothelial plasminogen activator inhibitor, is a protein that in Mouses is encoded by the SERPINE1 gene. PAI-1 is a serine protease inhibitor (serpin) that functions as the principal inhibitor of tissue plasminogen activator (tPA) and urokinase (uPA), the activators of plasminogen and hence fibrinolysis (the physiological breakdown of blood clots). It is a serine protease inhibitor (serpin) protein (SERPINE1). The other PAI, plasminogen activator inhibitor-2 (PAI-2) is secreted by the placenta and only present in significant amounts during pregnancy. In addition, protease nexin acts as an inhibitor of tPA and urokinase. PAI-1, however, is the main inhibitor of the plasminogen activators. PAI-1 is mainly produced by the endothelium (cells lining blood vessels), but is also secreted by other tissue types, such as adipose tissue.

PAI-1 inhibits the serine proteases tPA and uPA/urokinase, and hence is an inhibitor of fibrinolysis, the physiological process that degrades blood clots (1). PAI-1 inhibits the activity of matrix metalloproteinases, which play a crucial role in invasion of malignant cells across the basal lamina (2). Congenital deficiency of PAI-1 has been reported; as fibrinolysis is not suppressed adequately, it leads to a hemorrhagic diathesis (a tendency to hemorrhage). PAI-1 is present in increased levels in various disease states (such as a number of forms of cancer), as well as in obesity and the metabolic syndrome. It has been linked to the increased occurrence of thrombosis in patients with these conditions. In inflammatory conditions in which fibrin is deposited in tissues, PAI-1 appears to play a significant role in the progression to fibrosis (pathological formation of connective tissue). Presumably, lower PAI levels would lead to less suppression of fibrinolysis and conversely a more rapid degradation of the fibrin. Angiotensin II increases synthesis of PAI-1, so it accelerates the development of atherosclerosis.

References

1. Elokdah H, et al. (2004). J. Med. Chem. 47 (14): 3491–4.
2. Boncela, J, et al. (2001). J. Biol. Chem. (United States) 276 (38): 35305–11.

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