Endostatin is a naturally-occurring 20kD C-terminal fragment derived from type XVIII collagen. It is reported to serve as an anti-angiogenic agent, similar to angiostatin and thrombospondin. And It is produced by proteolytic cleavage of collagen XVIII, a member of the multiplexin family that is characterized by interruptions in the triple helix creating multiple domains, by proteases such as cathepsins. Using a genomic clone as a probe for fluorescence in situ hybridization, Endostatin was mapped the COL18A1 gene to 21q22.3. By immunoprecipitation analysis using membrane fractions of human mammary epithelial cells, It showed that endostatin specifically bound to cell surface nucleolin with high affinity. Blockage of nucleolin with neutralizing antibody or knockdown of nucleolin by RNA interference countered the antiendothelial activity of endostatin and abrogated its antiangiogenic and antitumor activity in vivo. Sample Type: Cell lysates, cell culture supernates, serum, plasma (heparin or EDTA). Intended Use: Sandwich High Sensitivity BioAssay(TM) ELISA kit for Quantitative Detection of Rat Endostatin. Sensitivity: <15pg/ml Range: 156pg/ml-10,000pg/ml Specificity: Recognizes natural and recombinant Rat Endostatin. There is no detectable cross-reactivity with other relevant proteins. Test Principle: This Rat Endostatin BioAssay(TM) ELISA Kit was based on standard sandwich enzyme-linked immune-sorbent assay technology. A specific for Endostatin has been precoated onto 96-well plates. Standards(Expression system for standard: NSO, Immunogen sequence: H1591-K1774) and test samples are added to the wells, a biotinylated detection specific for Endostatin is added subsequently and then followed by washing with PBS or TBS buffer. Avidin-Biotin-Peroxidase Complex was added and unbound conjugates were washed away with PBS or TBS buffer. HRP substrate TMB was used to visualize HRP enzymatic reaction. TMB was catalyzed by HRP to produce a blue color product that changed into yellow after adding acidic stop solution. The density of yellow is proportional to the Rat Endostatin amount of sample captured in plate. Kit Components: 96-well plate precoated with anti-Rat Col18a1 antibody, 1x plate Lyophilized recombinant Rat Col18a1 standard, 2x10ng/tube Biotinylated anti-Rat Col18a1 antibody, 1x130ul (dilution 1:100) Avidin-Biotin-Peroxidase Complex (ABC), 1x130ul (dilution 1:100) Sample diluent buffer, 1x30ml Antibody diluent buffer, 1x12ml ABC diluent buffer, 1x12ml TMB color developing agent, 1x10ml TMB stop solution, 1x10ml PBS washing buffer Powder for 1000ml, 1x buffer Storage and Stability: Store powder at 4C liquid at -20C. Store other components at 4C. Stable for at least 6 months For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.
