Elh mcp 1

Macrophages were treated with HCV core protein (10 μg/mL) for 24 h. Cell culture supernatants were collected and stored at -80°C in refrigerator. The amounts of CCL2 and CXCL10 present in culture supernatants were determined using specific Human CCL2 (ELH-MCP-1, Raybiotech) and Human CXCL10 (ELH-IP10, Raybiotech), Mouse CCL2 (EK287, MultiSciences) and Mouse CXCL10 (EMC121, NeobioScience) ELISA kit. The ELISA experiments were performed according to the manufacturer’s protocol.

To measure the protein level (pg/mL) of the chemokine C-C Motif Ligand 2 (CCL2, also known as human monocyte chemoattractant protein-1, MCP-1), we used an ELISA kit (Ray Biotech, ELH-MCP1) for this particular cytokine. Briefly, the standard curve for CCL2 was prepared side-by-side with supernatant representing different treatments. In a time-controlled sequence, the antibody pre-coated 96-well ELISA microplates were exposed to the following: a 100 µL of each sample/standard for 2.5 h, a 100 µL freshly diluted biotinylated antibody for an hour, a 100 µL Streptavidin solution for 45 min, and a 100 µL of the substrate reagent. Lastly, 50 µL of a stop-solution was added to each well to terminate the reaction. The intensity of CCL2 in the standard and each sample was measured at 450 nm using the Synergy HTX Multi-Reader (BioTek). The assay was repeated 3 times.