Nori Bovine P-Selectin ELISA Kit Summary

Alternative names for P-Selectin: SELP, CD62P, GMP-140, LECAM3, PADGEM

Alternative names for bovine: Cattle, cow, bull, buffalo,

P42201

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: 

P-selectin is encoded by the SELP gene.^[1] P-selectin functions as a cell adhesion molecule (CAM) on the surfaces of activated endothelial cells, which line the inner surface of blood vessels, and activated platelets. In unactivated endothelial cells, it is stored in granules called Weibel-Palade bodies. In unactivated platelets P-selectin is stored in α-granules. P-selectin is constitutively expressed in megakaryocytes and endothelial cells.^[2] P-selectin expression is induced by two distinct mechanisms. First, P-selectin is synthesized by megakaryocytes and endothelial cells, where it is sorted into the membranes of secretory granules.^[3] When megakaryocytes and endothelial cells are activated by agonists such as thrombin, P-selectin is rapidly translocated to the plasma membrane from granules.^[4] Secondly, increased levels of P-selectin mRNA and protein are induced by inflammatory mediators such as tumor necrosis factor-a (TNF-a), LPS, and interleukin-4 (IL-4). In ischemic stroke patients, plasma P-selectin concentration was reported to be highly correlated to plasminogen activator inhibitor-1 activity and tissue plasminogen activator activity.^[5 The primary ligand for P-selectin is P-selectin glycoprotein ligand-1 (PSGL-1) which is expressed on almost all leukocytes, although P-selectin also binds to heparan sulfate and fucoidan. P-selectin plays an essential role in the initial recruitment of leukocytes to the site of injury during inflammation. P-selectin is also very important in the recruitment and aggregation of platelets at areas of vascular injury. P-selectin has a functional role in metastasis of tumor similar to E-selectin.^[6] P-selectin helps cancer cells invade into bloodstream for metastasis and provided locally with multiple growth factors respectively.^[7]

References

1. Ryan US, et al. (1992). Opin. Immunol. 4 (1): 33–7.
2. Pan J, et al. (1998). Biol. Chem. 273 (16): 10058–67.
3. Disdier M, et al. (1992). Biol. Cell 3 (3): 309–21.
4. Hattori R, et al (1989). Biol. Chem. 264 (14): 7768–71.
5. Wang J, et al. (2005) Neurosci Lett. 384 (3): 305–9.
6. Köhler S, et al. (2010). J. Cancer 102 (3): 602–9.
7. Chen M, et al. (2006). Immunol. Ther. Exp. (Warsz.) 54 (2): 75–84.