Solution Information | help | |
Enzyme: | Interleukin-8 | |
inhibitor: | BDBM62334 | |
substrate: | n/a | |
Solution Type: | Aqueous | |
pH at Preparation: | n/a | |
Temp. Prep.: | n/a | |
Comments: | Protocol: 1. MCF7 cells stably overexpressing NOD1 2. GM-Tri-DAP (Sigma) 3. IL-8 OptEIA ELISA set (BD Biosciences, San Diego, CA) 4. SpectraMax 190 spectrophotometer (Molecular Devices) Protocol: 1. Compounds were diluted in DMEM 10% FBS plus Pen/Strep (culture medium), and 10 uL were added into respective wells (96-well culture plate) to reach desired final concentrations. 2. GM-Tri-DAP was added, in a final concentration of 5.4 ug/ml, to a culture medium suspension of MCF-7 cells (3,6x10+5 cells per mililiter). 3. Pre-induced MCF-7 cells were added into the respective wells (140 ul per well) and kept at 37oC (5% CO2 incubator) for 24 hours. 4. Thirty microliters of culture medium was used to measure IL-8 production using an IL-8 ELISA kit from BD according to the manufacturer's protocol. Briefly, 96-well ELISA plates (BD Biosciences) were coated with 100 uL per well of Capture Antibody diluted in Coating Buffer (100 mM Sodium carbonate, pH9.5) for overnight at 4oC. The well | |
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