Human mmp 2 quantikine elisa kit

Sample preparation: Cells (2×10⁶) were added to each 10 cm dish and cultured for 24 hours. To remove the unbound cells, the dishes were washed three times with serum-free medium. Next, the cells were incubated with 15 ml of serum-free media, and the media was collected using an Amicon Ultra 15 ml 10K column (Millipore) after 24 hours. uPA activity, MMP-2 and Cathepsin B concentrations were quantified using a uPA Activity Assay Kit (Millipore), a Human MMP-2 Quantikine ELISA Kit (R&D Systems) and a Human Cathepsin B Duo Set Kit (R&D Systems) according to the manufacturers recommended protocols. All of the sample analyses were performed in triplicate and repeated a minimum of three times.

Human MMP-2 Quantikine ELISA kit (R&D Systems, USA) and human MMP-9 ELISA kit (Abnova, Taiwan) were applied to test the expression of MMP-2 and MMP-9 respectively. After transfection, cells were incubated with fresh DMEM for 24 h, after which the culture medium was collected and centrifuged at 2000 rpm. Clarified medium was analyzed with ELISA kit (Institut Pourquier).

Commercially available diagnostic kits, Human MMP-2 Quantikine ELISA Kit and Human MMP-9 Quantikine Immunoassay (R&D System, Abingdon, UK), were applied to gelatin concentration evaluation. The gelatinase levels were expressed in nanograms per milliliter in plasma and in nanograms per milligram of protein in vein wall homogenates. VEGF concentration was determined using diagnostic kit Human VEGF Quantikine Immunoassay (R&D System, Abingdon, UK) and expressed in picograms per milliliter in plasma, and nanograms per gram and nanograms per milligram of protein in tissue. Protein level in tissue samples was estimated with the usage of commercially available Bradford reagent (BIO-RAD Protein Assay, Hercules, USA). The assays were performed with the usage of Thermoshaker DTS-4 (ELMI, Calabasas, North America) and Microplate Reader Model 680 (BIO-RAD, Hercules, USA) with software Microplate Manager version 5.2.1 (BIO-RAD, Hercules, USA). All measurements were performed according to manufacturers’ manuals.