TOPFlash Assay Wnt/β-catenin HEK293 Reporter Cell Lines

Product Data:

Figure 1. EGFP activation in TCF/LEF1 HEK293 Reporter Cells following Wnt pathway stimulation.
TOPFlash TCF/LEF1 (EGFP) HEK293 cells (RCL-0003-5S) were seeded at 5 × 10⁴ cells per well in a 48-well plate and incubated for 8–10 hours. Cells were treated with 2.5 mM LiCl for 16 hours, followed by the addition of WNT3a (25 ng/ml) and incubation for an additional 16 hours. EGFP expression was visualized using fluorescence microscopy and quantified by flow cytometry.

Figure 2. Fluorescence detection of Wnt/β-catenin activation in TCF/LEF1 HEK293 Reporter Cells

TOPFlash TCF/LEF1 (EGFP) HEK293 cells (RCL-0003-5S) were seeded at 5 × 10⁴ cells per well in a 48-well plate and incubated for 8–10 hours. Cells were treated with LiCl at the indicated concentrations for 16 hours. WNT3a was then added to a final concentration of 25 ng/ml, followed by an additional 16-hour incubation. EGFP expression was visualized using fluorescence microscopy and analyzed by flow cytometry after trypsinization.

Figure 3. Luminescence detection of Wnt/β-catenin activation in TCF/LEF1 HEK293 Reporter Cells.
TOPFlash TCF/LEF1 Firefly Luciferase Reporter HEK293 Cells (RCL-0003-3S) were treated with LiCl at the indicated concentrations for 36 hours. Following stimulation, cells were lysed using LipExoGen lysis buffer (DTB-001E). Supernatants were transferred to a 96-well plate containing LipExoGen’s Firefly Assay Buffer (DTB-001B) and Firefly Substrate (DTB-001A), and luminescence was measured. Firefly luciferase signal intensity increased proportionally with LiCl concentration, reflecting Wnt/β-catenin pathway activation.

Key Features:

• Reporter system:
  • Fluorescent or luminescent reporters enable real-time detection of Wnt/β-catenin signaling in live cells.
• Stably integrated TCF/LEF1-responsive promoter driving reporter expression.
• Validated with Wnt3a and/or LiCl to confirm canonical Wnt pathway responsiveness.
• Ideal for live-cell imaging, flow cytometry, or luminescence-based assays depending on format.

Applications:

• Screening Wnt pathway activators and inhibitors.
• Studying TCF/LEF1 transcriptional activity in mammalian systems.
• Evaluating β-catenin-mediated transcriptional regulation.
• Fluorescence or luminescence-based readouts for high-throughput screening and mechanistic studies.

Quality Control: Each cell line is validated for reporter expression and Wnt responsiveness using Wnt3a and LiCl stimulation. Stable expression is confirmed for ≥20 passages. All cells are screened and certified mycoplasma-free.

Shipping and Storage: Cells are shipped on dry ice and should be stored in the vapor phase of liquid nitrogen upon receipt.

How to Use: Plate cells and stimulate with Wnt3a or LiCl. Monitor reporter activity via fluorescence microscopy, flow cytometry, or luminescence detection depending on the construct.

Contents:

• 1 vial containing >1 × 10⁶ Wnt/β-catenin HEK293 Reporter Cells (variant-specific).

Materials Required but Not Supplied:

• Media for Cell Culture:
  • Thaw Medium: DMEM with 10% FBS and 1% Penicillin/Streptomycin.
  • Growth Medium: DMEM with 10% FBS and appropriate antibiotic (e.g., 1 µg/ml Puromycin or 10 µg/ml Blasticidin).
• Materials for Reporter Assays:
  • Recombinant Wnt3a or LiCl
  • 96-well tissue culture-treated white clear-bottom assay plate
  • Fluorescence microscope, flow cytometer, or luminometer

Background:

Wnt/β-catenin signaling plays a pivotal role in development, stem cell regulation, and disease processes such as cancer. Activation of the canonical pathway leads to β-catenin stabilization and nuclear translocation, where it binds TCF/LEF transcription factors to induce target gene expression. The TOPFlash system leverages multimerized TCF/LEF binding elements to drive reporter expression, enabling sensitive and specific measurement of pathway activation. These HEK293 reporter lines offer robust tools for dissecting Wnt-driven transcriptional responses and screening pathway modulators in live cells.

These products are covered by a Limited Use License. For more information, please email info@lipexogen.com