Nori Canine E-Selectin ELISA Kit Summary

Alternative names for E-Selectin: SELE, ELAM1, LECAM2, CD62E

Alternative name for canine: Dog

P33730

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: 

E-selectin is encoded by the SELE gene. Different from P-selectin, which is stored in vesicles called Weibel-Palade bodies, E-selectin is not stored in the cell and has to be transcribed, translated, and transported to the cell surface. The production of E-selectin is stimulated by the expression of P-selectin which in turn, is stimulated by tumor necrosis factor α (TNFα), interleukin-1 (IL-1) and lipopolysaccharide (LPS).^[1] It takes about two hours, after cytokine recognition, for E-selectin to be expressed on the endothelial cell’s surface. Maximal expression of E-selectin occurs around 6–12 hours after cytokine stimulation, and levels returns to baseline within 24 hours.^[1] Shear forces are also found to affect E-selectin expression. A high laminar shear enhances acute endothelial cell response to interleukin-1β in naïve or shear-conditioned endothelial cells as may be found in the pathological setting of ischemia/reperfusion injury while conferring rapid E-selectin down regulation to protect against chronic inflammation.^[2] E-selectin recognizes and binds to sialylated carbohydrates present on the surface proteins of certain leukocytes. E-selectin ligands are expressed by neutrophils, monocytes, eosinophils, memory-effector T-like lymphocytes, and natural killer cells. Each of these cell types is found in acute and chronic inflammatory sites in association with expression of E-selectin, thus implicating E-selectin in the recruitment of these cells to such inflammatory sites. During inflammation, E-selectin plays an important part in recruiting leukocytes to the site of injury. Cancer cells are able to infiltrate the inflammatory system by interacting with selectins. E-selectin mediates the adhesion of tumor cells to endothelial cells, by binding to E-selectin ligands. The immunohistochemical expressions of E-selectin and PECAM-1 were significantly increased at intima in vulnerable plaques of acute coronary syndrome (ACS) group, especially in neovascular endothelial cells, and positively correlated with inflammatory cell density, suggesting that PECAM-1 and E-selectin might play an important role in inflammatory reaction and development of vulnerable plaque. E-selectin Ser128Arg polymorphism is associated with ACS, and it might be a risk factor for ACS.^[24]

References

1. Leeuwenberg JF, et al. (1992) Immunology 77 (4): 543–9.
2. Huang RB, et al. (2012) PLoS ONE 7 (2): e31874.
3. Fang F, et al. (2011) Zhonghua Xin Xue Guan Bing Za Zhi (in Chinese) 39 (12): 1110–6.