NF-kB Luciferase Stable RAW264.7 Cell Line
|T3015||1x106 cells / 1.0 ml|
|Description||Luciferase is commonly used as a reporter to assess the transcriptional activity of a genetic construct with luciferase gene under the promoter of interest. NF-kB Luciferase Stable RAW264.7 Cell Line has been stably transfected with a 3kB-Luc SV40 reporter, which contains 3 NF-kB sites from the interferon gene upstream of the luciferase coding region. This cell line is useful in investigation of NF-kB activation in monocyte-macrophage models. This cell line should be maintained in 100ug/ml G418.|
|Species||Mouse (M. musculus)|
|Species description||Mus musculus|
|Seeding Density||50,000 - 70,000 cells/cm2. Recommended split ratio of 1:5|
|Population Doubling Time||30 - 35 hours|
For Research Use Only
|Unit quantity||1x106 cells / 1.0 ml|
|Pharmaceutical Target||Catalytic Receptors|
|Caution||For Research Use Only|
|Cell Type||Drug Discovery Cells|
|Propagation Requirements||The base medium for this cell line is Prigrow III medium available at abm, Cat. No. TM003. To make the complete growth medium, add the following components to the base medium: fetal bovine serum (TM999) to a final concentration of 10%, L-Glutamine to 2mM, 100ug/ml G418, and Penicillin/Streptomycin Solution (G255) to a final concentration of 1%. Carbon dioxide (CO2): 5%, Temperature: 37.0°C.|
|Subculture Protocol||1. Subculture when the culture reaches 80-85% confluency or above.|
2. Warm complete medium, Trypsin-EDTA solution, and DPBS (Ca and Mg free) to room temperature.
3. Rinse the cells with DPBS.
4. Add 2-3 ml of Trypsin-EDTA solution into flask. Gently rock the flask to ensure complete coverage of cells by T/E solution. Incubate the flask in a 37
Have been tested for luciferase activity
1. For for-profit organizations and corporations, please contact [email protected] for pricing of this item.
2. Sale of this item is subjected to the completion of a Material Transfer Agreement (MTA) by the purchasing individual/institution for each order. If you have any questions regarding this, please contact us at [email protected].
3. All test parameters provided in the CoA are conducted using abm's standardized culture system and procedures. The stated values may vary under the end-user's culture conditions. Please verify that the product is suitable for your studies by referencing published papers or ordering RNA (0.5 μg, Cat.# C207, $450.00) or cell lysate (100 μg, Cat.# C206, $600.00) to perform preliminary experiments, or alternatively use our Gene Expression Assay Service (Cat# C138). All sales are final.
4. We recommend live cell shipments for ease of cell transfer and this option can be requested at the time of ordering. Please note that the end-user will need to evaluate the feasibility of live cell shipment by taking into account the final destination's temperature variation and its geographical location. In addition, we thoroughly test our cell lines for freeze-thaw recovery. If frozen cells were received and not recovered in your lab under the exact, specified conditions (using recommended culture vessel, media, additional supplements, and atmospheric conditions), a live cell replacement is possible at a cost (plus shipping).
5. All of abm's cell biology products are for research use ONLY and NOT for therapeutic/diagnostic applications. abm is not liable for any repercussions arising from the use of its cell biology product(s) in therapeutic/diagnostic application(s). Please contact a technical service representative for more information.
6. abm makes no warranties or representations as to the accuracy of the information on this site. Citations from literature and provided for informational purposes only. abm does not warrant that such information has been shown to be accurate.
7. abm warrants that cell lines shall be viable upon initiation of culture for a period of thirty (30) days after shipment and that they shall meet the specifications on the applicable abm Material Product Information sheet, certificate of analysis, and/or catalog description. Such thirty (30) day period is referred to herein as the "Warranty Period."
|Depositor||University of Western Australia|
|Is the level of NFkB activationby RANK Ligand passage dependent? How many passages can I obtain the same (high) level of the NFkB activation by RANK ligand?|
No major difference was observed in the level of the NFkB activation within passage 5. No comparative experiments have been run after this time line to test further.
|From which species is the luciferase gene expressed in the NF-kB Luciferase Stable RAW264.7 Cell Line? Is it the firefly luciferase gene or the sea pansy luciferase gene?|
This cell line expresses Firefly (Photinus pyralis) luciferase.
|How was luciferase activity assayed?|
The luciferase assay was performed using our Luciferase Assay Kit (Cat# G287) and the protocol can be found on that webpage.
|Does this cell line consistently express firefly luciferase or does it need to be incubated with RANKL?|
These cells need RANKL stimulation for luciferase reporter gene expression as it is NF-κB-driven luciferase reporter gene and RANKL stimulates NF-κB.
|How much Pen/Strep is needed?|
You may consider Cat# G255 Pen/Strep. Penicillin- Streptomycin contains 10,000 units of penicillin (base), 10,000µg of streptomycin (base) per mL in WFI water. It is supplied as a 0.22 µm-filtered, 100X frozen liquid. As a 100X concentration, it is designed to be supplemented as 1 ml per 99 ml of basal medium, yielding an iso-osmotic solution at final dilution (i.e. this is 100X and needs to be used as 1X final).
|Does T3015 contain any antibiotic resistance?|
Yes, the selection marker for this cell line is G418.
|What concentration of G418 is required to maintain the T3015 cell line?|
100ug/ml of G418 is required for maintenance of the cell line
- Li, X et al. "Sinomenine suppresses osteoclast formation and Mycobacterium tuberculosis H37Ra-induced bone loss by modulating RANKL signaling pathways" PLoS One 8(9):e74274 (2013). DOI: 10.1371/journal.pone.0074274.
- Singh, PP et al. "Membrane-bound receptor activator of NFκB ligand (RANKL) activity displayed by osteoblasts is differentially regulated by osteolytic factors" Biochem Biophys Res Commun 422(1):48-53 (2012). DOI: 10.1016/j.bbrc.2012.04.103.
- Liu, Q et al. "SC-514, a selective inhibitor of IKKβ attenuates RANKL-induced osteoclastogenesis and NF-κB activation" Biochem Pharmacol. 86(12):1775-83 (2013). DOI: 10.1016/j.bcp.2013.09.017. PubMed: 24091016.
- van der Kraan, AG et al. "HSP90 inhibitors enhance differentiation and MITF (microphthalmia transcription factor) activity in osteoclast progenitors" Biochem J. 451(2):235-44 (2013). DOI: 10.1042/BJ20121626. PubMed: 23379601.
- King, TJ et al. "Methotrexate chemotherapy promotes osteoclast formation in the long bone of rats via increased pro-inflammatory cytokines and enhanced NF-κB activation" Am J Pathol. 181(1):121-9 (2012). DOI: 10.1016/j.ajpath.2012.03.037. PubMed: 22642908.
- Wang, C et al. "12-O-tetradecanoylphorbol-13-acetate (TPA) inhibits osteoclastogenesis by suppressing RANKL-induced NF-kB activation" J Bone Miner Res 18:2159-2168 (2003). DOI: 10.1359/jbmr.2003.18.12.2159.
- Xu, J et al. "Evidence of reciprocal regulation between the high extracellular calcium and RANKL signal transduction pathways in RAW cell derived osteoclasts" J Cell Physiol. 202(2):554-62 (2005). PubMed: 15389575.
- de Camargo, A. C., Biasoto, A. C. T., Schwember, A. R., Granato, D., Rasera, G. B., Franchin, M., ... & Shahidi, F. "Should we ban total phenolics and antioxidant screening methods? The link between antioxidant potential and activation of NF-κB using phenolic compounds from grape by-products" Food chemistry 290:229-238 (2019).