TLR4/IL-8 cell line

TLR4/IL-8 Leeporter Luciferase Reporter-HeLa Cell Line

Cat. Num. AAA669524

• Applications: Functional Assay
• Synonyms: TLR4/IL-8; TLR4/IL-8 Leeporter Luciferase Reporter-HeLa Cell Line; TLR4/IL-8 cell line

• Host: HeLa cells
• Form/Format: Each vial contains 2-3 x 10^6 cells in 1 ml of 90% FBS + 10% DMSO.

• Applicable Applications for TLR4/IL-8 cell line: Functional Assay (FA)
• Application Notes: Monitor the TLR4 signaling pathway activity. Screen for activators or inhibitors of the TLR4 signaling pathway.
• Culture Conditions: Cells should be grown at 37 degree C with 5% CO2 using DMEM medium (w/ L-Glutamine, 4.5g/L Glucose and Sodium Pyruvate) supplemented with 10% heat-inactivated FBS and 1% Pen/Strep, plus 1ug/ml of Puromycin, 5ug/ml Blasticidin and 500ug/ml G418 (Note: Puromycin, Blasticidin and G418 can be omitted during the reporter cell assays). It is recommended to quickly thaw the frozen cells upon receipt or from liquid nitrogen in a 37 degree C water-bath, transfer to a tube containing 10ml of growth medium without Puromycin, Blasticidin and G418, spin down cells, resuspend cells in pre-warmed growth medium without Puromycin, Blasticidin and G418, transfer resuspended cells to T25 flask and culture in 37 degree C-CO2 incubator. Leave the T25 flask in the incubator for 1~3 days without disturbing or changing the medium until cells completely recover viability and become adherent. Once cells are over 90% adherent, remove growth medium and passage the cells through trypsinization and centrifugation. At first passage, switch to growth medium containing Puromycin, Blasticidin and G418. Cells should be split before they reach complete confluence. To passage the cells, detach cells from culture vessel with Trypsin/EDTA, add complete growth medium and transfer to a tube, spin down cells, resuspend cells and seed appropriate aliquots of cells suspension into new culture vessels. Subcultivation ration = 1:10 to 1:20 weekly. To achieve satisfactory results, cells should not be passaged over 16 times
• Functional validation: A. Response of TLR4/IL-8 Leeporter - HeLa cells to LPS.; 1. Harvest TLR4/IL-8 Leeporter - HeLa cells and seed cells into a white solid-bottom 96-well microplate in 100 ul of growth medium at 5 x 10^4 cells/well.; 2. Incubate cells at 37 degree C in a CO2 incubator for overnight.; 3. The next day, stimulate cells with various concentrations of LPS.; 4. Incubate at 37 degree C in a CO2 incubator for 6-16 hours.; 5. Add 50 ul of luciferase assay reagent (Cat #MBS668917) per well.; 6. Incubate at room temperature for 1-5 minutes and measure luminescence using a microplate luminometer.
• Shipping Note: Product available only in the USA.
• Dry Ice Shipment: Extra charge fee may add to your shipping cost as dry ice is required to ship this product.
• Preparation and Storage: Immediately upon receipt, store in liquid nitrogen.

Activity

(Fig-1: Induction of TLR4 activity by LPS in TLR4/IL-8 Leeporter™ – HeLa cells.)

Related Product Information for TLR4/IL-8 cell line

The TLR4/IL-8 Leeporter Luciferase Reporter cell line is a stably transfected HeLa cell line which expresses human TLR4, MD-2 and CD14 as well Renilla luciferase reporter gene under the transcriptional control of the IL-8 promoter. IL-8 is one of the major pro-inflammatory cytokines induced by ligand (such as LPS)-mediated Toll-like receptor 4 (TLR4) activation. TLR4 is one of the key innate immune receptors, which is activated by LPS and can lead to sepsis upon dysregulation. The TLR4/IL-8 activation by LPS is shown in Figure 1.

Product Categories/Family for TLR4/IL-8 cell line

Reporter Cell Lines