Active Heparanase (HPSE)

Heparanase (HPSE) selectively cleaves heparan sulfate at specific sites on heparan sulfate proteoglycans (HSPGs). HPSE facilitates cell migration associated with metastasis, wound healing and inflammation. An increase in its activity is associated with an increase in VEGF activity, which further enhances angiogenesis. HPSE also enhances shedding of syndecans and increases endothelial invasion and angiogenesis in myelomas. It acts as a procoagulant by increasing the generation of activation factor X in the presence of tissue factor and activation factor VII. In addition, it increases cell adhesion to the extracellular matrix (ECM), independent of its enzymatic activity. HPSE is highly expressed in placenta and spleen and weakly expressed in lymph node, thymus, peripheral blood leukocytes, bone marrow, endothelial cells, fetal liver and tumor tissues. The enzyme activity of recombinant human HPSE was assayed in an ELISA format using non-reducing end biotinylated heparan sulfate on recombinant human syndecan 4 as a substrate. Briefly, combining 10 µL of different concentrations of rhHPSE with 10 µL of the biotinylated-syndecan 4 in a vial (10 µL of assay buffer and 10 µL of biotinylated-syndecan 4 as a negative control) and incubated for 2h at 37℃. After incubation, 220 µL of PBS, with 1% BSA (pH7.4) was added to each reaction, then loading 100 ul of each sample onto the anti-syndecan 4 pAb-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST 3 times and incubation with Streptavidin-HRP for 30min, then wells were aspirated and washed 5 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50 µl stop solution to the wells and read at 450nm immediately. As a result, 500 ng of recombinant human HPSE digestion will result in >50% of OD reduction compared with the negative control.