DESCRIPTION

E-selectin, also known as ELAM-1 or CD62E, is a member of the selectin family of adhesion molecules that also include L-selectin and P-selectin. E-selectin is an inducible endothelial cell surface molecule. Its expression on endothelial cells is transcriptionally upregulated by various proinflammatory substances such as IL-1, TNFα and lipopolysaccharide (LPS). Besides, it can be secreted by endothelial cells, and can be detected in a soluble form (sE-selectin) in serum. During inflammation, Eselectin plays an important part in recruiting leukocytes to the site of injury.

PRINCIPLE OF THE ASSAY

An anti-tag antibody is pre-coated onto the bottom of wells. After adding antigen or samples, Tagged-Capture antibody and HRP-conjugated detection antibody, a sandwich complex is formed in the solution. TMB acts as a HRP substrate, and the solution color will change from colorless to blue. A stop solution containing sulfuric acid turns the solution yellow. The color intensity is proportional to the quantity of bound protein, which is measurable at 450 nm with the correction wavelength set at 630 nm.

PRECISION

Intra-assay Precision (Precision within an assay) Three samples of known concentration were tested 8 times on one plate to assess intra-assay precision. Inter-assay Precision (Precision between assays) Three samples of known concentration were tested in 16 separate assays to assess inter-assay precision.

Intra-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	8	4,545.4	247.2	5.4
2	8	1,187.7	55.0	4.6
3	8	686.5	12.4	1.8

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	16	4,569.9	243.5	5.3
2	16	1,179.9	50.8	4.3
3	16	683.2	19.3	2.8

RECOVERY

The recovery of mouse E-selectin/CD62E spiked to three different levels throughout the range of the assay in mouse serum was evaluated.

Sample Type		Average% of Expected	Range (%)
mouse serum	1:160	100	95-110
	1:320	103	80-116

LINEARITY

To assess the linearity of the assay, samples were diluted with the appropriate Sample Diluent to produce samples with values within the dynamic range of the assay. (The mouse serum was initially diluted 1:20.)

		Mouse serum
1:2	Average% of Expected	100
	Range (%)	-
1:4	Average% of Expected	124
	Range (%)	118-131
1:8	Average% of Expected	125
	Range (%)	117-133
1:16	Average% of Expected	110
	Range (%)	107-113

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human Mouse E-selectin/CD62E One-Step ELISA Kit produced at Sunlong Medical™.