Human ESM1 protein, His tag

Endothelial cell-specific molecule 1 (ESM1) is a proteoglycan secreted by endothelial cells (primarily in the human lung and kidney tissues) and its mRNA expression is regulated by inflammatory cytokines. Endocan expression, which detected in various epithelia and adipocytes has been shown to be upregulated by vascular endothelial growth factor (VEGF), fibroblast growth factor 2 (FGF-2), TNF alpha, IL1 beta, or lipopolysaccharide and downregulated by IFN gamma. Genetically engineered cells overexpressing ESM1 induce tumor formation, implying that ESM1 might be involved in the pathophysiology of tumor growth in vivo. Besides, Lymphocyte Function Associated Antigen 1 Alpha (LFA1a) has been identified as an interactor of ESM1, thus a binding ELISA assay was conducted to detect the interaction of recombinant human ESM1 and recombinant human LFA1a. Briefly, ESM1 were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to LFA1a-coated microtiter wells and incubated for 2h at 37ºC. Wells were washed with PBST and incubated for 1h with anti- ESM1 pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody, wells were aspirated and washed 3 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37ºC. Finally, add 50 μl stop solution to the wells and read at 450nm immediately. The binding activity of of ESM1 and LFA1a was in a dose dependent manner.