Clear bottom 96-well plates were seeded with 5×103 MCF-7 cells per well and treated with 0, 1, 10, or 100 ng/mL purified recombinant cmvIL-10 (R&D Systems) in triplicate. Cell supernatants were harvested at various time points for subsequent analysis. Cell lysates were also collected from these plates by adding 100μL cell lysis buffer (150mM NaCl, 20 mM HEPES, 0.5% Triton X-100, 1mM NaOV4, 1mM EDTA, 0.1% NaN3) with fresh protease inhibitor (Calbiochem, La Jolla, CA) onto the cells and administering a freeze-thaw cycle. MMP-10 protein levels were determined by sandwich ELISA according to manufacturer’s instructions (R&D Systems Duo Set DY910). Protein concentrations were determined by interpolation from a standard curve with an R2 value greater than 0.99 using the Opsys MR plate reader and Revelation Quicklink 4.24 software (Dynex Technologies, Chantilly, VA).
Purified recombinant cmvil 10
Manufactured by R&D Systems
Purified recombinant cmvIL-10 is a laboratory research product. It is a cytokine protein expressed in a recombinant system. The protein's core function is to support in vitro studies.
Automatically generated - may contain errors
Lab products found in correlation
Opsys mr plate reader, by Dynex (1 mentions)
Stat3 inhibitor s3i 201, by Santa Cruz Biotechnology (1 mentions)
Epidermal growth factor (egf), by R&D Systems (1 mentions)
Hff cell line, by American Type Culture Collection (1 mentions)
Human il 10, by R&D Systems (1 mentions)
Mda mb 231 human breast adenocarcinoma cells, by American Type Culture Collection (1 mentions)
2 protocols using purified recombinant cmvil 10
1
Quantification of MMP-10 Secretion in MCF-7 Cells
2
Breast Cancer and Cytomegalovirus Interactions
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MDA-MB-231 human breast adenocarcinoma cells (American Type Culture Collection, Manassas, VA) were cultured in L-15 Leibovitz’s Medium (Mediatech, Manassas, VA) supplemented with 10% fetal bovine serum (FBS) (Atlanta Biologicals, Flowery Branch, GA) at 37 °C with atmospheric CO2. The human foreskin fibroblast (HFF) cell line (ATCC) was cultured in Dulbecco’s Modified Eagle Medium (DMEM) with 10% FBS at 37 °C in a humidified chamber with 5% CO2. Human cytomegalovirus strain AD169 (ATCC) was used to infect confluent monolayers of HFFs at the indicated multiplicities of infection. Purified recombinant cmvIL-10, human IL-10, and epidermal growth factor (EGF) were purchased from R&D Systems (Minneapolis, MN). IL-10R neutralizing antibody and S3I-201 Stat3 inhibitor were from Santa Cruz Biotechnology (Santa Cruz, CA).
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