KC-0203

293T-Wnt-Luciferase-Reporter-Cell-Line

20005

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Background of 293T-Wnt-Luciferase-Reporter-Cell-Line

NFκB signaling plays a critical role in developmental process, including cell proliferation, differentiation and tissue patterning, and can be aberrantly activated in cancer and diabetes. When Wnt pathway is activated, the β-catenin protein complex translocate from cytoplasm to nucleus, regulates a wide spectrum of gene expression after binds to its response element on the promoter region of downstream gene.

Specifications

Catalog Number	KC-0203
Cell Line Name	293T-Wnt-Luciferase-Reporter-Cell-Line
Host Cell Line	293T
Description	HEK293T cell line stable expressing exogenous firefly luciferase gene under the control of TCF-responsive element
Quantity	Two vials of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Drug screening and biological assays
Freezing Medium	70% DMEM + 20% FBS + 10% DMSO
Propagation Medium	DMEM + 10% FBS
Selection Marker	NA
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:5 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 30 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

293T Wnt luciferase cell line was generated using plasmid transfection method.

Characterization

Figure: 293T Wnt luciferase cell line was seed into the 96-well plate, and treated with rmWnt3a for 16 hours, then readout with Bright-Glo system

Cell model for monitoring Wnt pathway.

Example: PORCN inhibitor test in co-culture reporter assay of 293-wnt-reporter line and L-wnt3a line

Cell Resuscitation

Prewarm culture medium (DMEM supplemented with 10% FBS)in a 37°C water bath.
Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
Incubate the flask at 37°C, 5% CO₂ incubator.
Split saturated culture 1:4-1:5 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% DMEM + 20% FBS + 10% DMSO) fresh immediately before use.
Keep the freezing medium on ice and label cryovials.
Transfer cells to a sterile, conical centrifuge tube, and count the cells.
Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
Aliquot 1 mL of the cell suspension into each cryovial.
Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
Transfer vials to liquid nitrogen for long-term storage

References

MacDonald, B. T., Tamai, K. & He, X. Wnt/β-Catenin Signaling: Components, Mechanisms, and Diseases. Dev.
Cell 17, 9–26 (2009).
Nile, A. H. & Hannoush, R. N. Fatty acylation of Wnt proteins. Nat. Chem. Biol. 12, 60–69 (2016).
Cheng, Y. et al. Wnt-C59 arrests stemness and suppresses growth of nasopharyngeal carcinoma in mice by
inhibiting the Wnt pathway in the tumor microenvironment. Oncotarget 6, 14428–14439 (2015).