HRG ELISA Kit (Human)

Intra-assay Precision (Precision within an assay) : 3 samples with low, middle and high level Histidine Rich Glycoprotein (HRG) were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays) : 3 samples with low, middle and high level Histidine Rich Glycoprotein (HRG) were tested on 3 different plates, 8 replicates in each plate. CV (%) = SD/meanX100 Intra-Assay: CV<10% Inter-Assay: CV<12%

Plasma glycoprotein that binds a number of ligands such as heme, heparin, heparan sulfate, thrombospondin, plasminogen, and divalent metal ions. Binds heparin and heparin/glycosaminoglycans in a zinc-dependent manner. Binds heparan sulfate on the surface of liver, lung, kidney and heart endothelial cells. Binds to N-sulfated polysaccharide chains on the surface of liver endothelial cells. Inhibits rosette formation. Acts as an adapter protein and is implicated in regulating many processes such as immune complex and pathogen clearance, cell chemotaxis, cell adhesion, angiogenesis, coagulation and fibrinolysis. Mediates clearance of necrotic cells through enhancing the phagocytosis of necrotic cells in a heparan sulfate-dependent pathway. This process can be regulated by the presence of certain HRG ligands such as heparin and zinc ions. Binds to IgG subclasses of immunoglobins containing kappa and lambda light chains with different affinities regulating their clearance and inhibiting the formation of insoluble immune complexes. Tethers plasminogen to the cell surface. Binds T-cells and alters the cell morphology. Modulates angiogenesis by blocking the CD6-mediated antiangiongenic effect of thrombospondins, THBS1 and THBS2. Acts as a regulator of the vascular endothelial growth factor (VEGF) signaling pathway; inhibits endothelial cell motility by reducing VEGF-induced complex formation between PXN/paxillin and ILK/integrin-linked protein kinase and by promoting inhibition of VEGF-induced tyrosine phosphorylation of focal adhesion kinases and alpha-actinins in endothelial cells. Also plays a role in the regulation of tumor angiogenesis and tumor immune surveillance. Normalizes tumor vessels and promotes antitumor immunity by polarizing tumor-associated macrophages, leading to decreased tumor growth and metastasis.

The test principle applied in this kit is Sandwich enzyme immunoassay. The microtiter plate provided in this kit has been pre-coated with an antibody specific to Histidine Rich Glycoprotein (HRG) . Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody specific to Histidine Rich Glycoprotein (HRG) . Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After TMB substrate solution is added, only those wells that contain Histidine Rich Glycoprotein (HRG), biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450nm +/- 10nm. The concentration of Histidine Rich Glycoprotein (HRG) in the samples is then determined by comparing the O.D. of the samples to the standard curve.

Reconstitution & Storage Instructions Western Blotting/Immunoblotting (WB/IB) Protocol Immunohistochemistry (IHC) Protocol Immunocytochemistry (ICC) Protocol Enzyme-Linked ImmunoSorbent Assay (ELISA) Protocol Blocking Peptide Competition Protocol (BPCP) Immunoprecipitation (IP) Protocol Antibody Array (AA) Protocol Reconstitution & Storage Instructions Reconstitution & Storage Instructions Western Blotting/Immunoblotting (WB/IB) Protocol Western Blotting/Immunoblotting (WB/IB) Protocol Immunohistochemistry (IHC) Protocol Immunohistochemistry (IHC) Protocol Immunocytochemistry (ICC) Protocol Immunocytochemistry (ICC) Protocol Enzyme-Linked ImmunoSorbent Assay (ELISA) Protocol Enzyme-Linked ImmunoSorbent Assay (ELISA) Protocol Blocking Peptide Competition Protocol (BPCP) Blocking Peptide Competition Protocol (BPCP) Immunoprecipitation (IP) Protocol Immunoprecipitation (IP) Protocol Antibody Array (AA) Protocol Antibody Array (AA) Protocol

Matrix Recovery Range (%) Average (%) Serum (n=5) 97-104 101 EDTA Plasma (n=5) 81-91 88 Heparin Plasma (n=5) 96-103 101 Matrix Recovery Range (%) Average (%) Matrix Recovery Range (%) Average (%) Serum (n=5) 97-104 101 Serum (n=5) 97-104 101 EDTA Plasma (n=5) 81-91 88 EDTA Plasma (n=5) 81-91 88 Heparin Plasma (n=5) 96-103 101 Heparin Plasma (n=5) 96-103 101

Component Amount Anti-HRG Microplate 96 Wells (12 x 8 Well Strips) HRG Lyophilized Standard 2 x 30 ng 100X Biotinylated HRG Detector Antibody 120 uL Avidin/HRP Conjugate 120 uL Standard Diluent 1 x 20 mL Detector Antibody Diluent 1 x 12 mL Conjugate Diluent 1 x 12 mL 30X Wash Buffer 1 x 20 mL TMB Substrate 1 x 9 mL Stop Solution 1 x 6 mL Component Amount Component Amount Anti-HRG Microplate 96 Wells (12 x 8 Well Strips) Anti-HRG Microplate 96 Wells (12 x 8 Well Strips) HRG Lyophilized Standard 2 x 30 ng HRG Lyophilized Standard 2 x 30 ng 100X Biotinylated HRG Detector Antibody 120 uL 100X Biotinylated HRG Detector Antibody 120 uL Avidin/HRP Conjugate 120 uL Avidin/HRP Conjugate 120 uL Standard Diluent 1 x 20 mL Standard Diluent 1 x 20 mL Detector Antibody Diluent 1 x 12 mL Detector Antibody Diluent 1 x 12 mL Conjugate Diluent 1 x 12 mL Conjugate Diluent 1 x 12 mL 30X Wash Buffer 1 x 20 mL 30X Wash Buffer 1 x 20 mL TMB Substrate 1 x 9 mL TMB Substrate 1 x 9 mL Stop Solution 1 x 6 mL Stop Solution 1 x 6 mL

This assay has high sensitivity and excellent specificity for detection of Histidine Rich Glycoprotein (HRG) . No significant cross-reactivity or interference between Histidine Rich Glycoprotein (HRG) and analogues was observed.