topflash mcherry reporter  (Addgene inc)

Journal: bioRxiv

Article Title: A synNotch-based morphogen detection system reveals sFRP2 enhances Wnt3a signaling

doi: 10.64898/2026.02.09.704138

Figure Lengend Snippet: (A) Schematic illustration of HEK293T cells expressing TOPFlash mCherry reporter. (B) TOPFlash reporter assay with recombinant sFRP2. Reporter cells were incubated with the conditioned media harvested from GFP-Wnt3a-secreting cells and supplemented with various concentrations of recombinant sFRP2. The conditioned media were applied either at full concentration (right) or diluted with an equivalent volume of DMEM (left). After 48-h incubation, the mCherry reporter activation was measured by flow cytometry. The potentiating effect of sFRP2 on Wnt signaling was pronounced under the diluted conditions (left). (C) Experimental setup for the gradient assay. GFP-Wnt3a-secreting L cells (1.6×10 4 cells) were seeded into the left chamber, while HEK293T TOPFlash reporter cells (1.6×10 4 cells) were seeded into the right chamber. After incubation overnight, the insert was removed, and the media were replaced with 1% agarose-containing media. Standard DMEM or DMEM containing recombinant sFRP2 was subsequently added over the solidified agarose gel. The confocal images were acquired at 24 h intervals. (D) Gradient patterns after 4 days of incubation. The spatial distribution of mCherry signals was extended in an sFRP2-dose-dependent manner. Red lines indicate the mean mCherry intensity derived from 5 subdivided vertical regions per image. Specifically, each 900-pixel vertical image was partitioned into 5 equal segments of 180 pixels each. The mean fluorescence intensity was calculated for each segment, and the red line represents the average of these five regional means. Shaded areas in the graph represent the ±SD. Scale bar: 200 µm.

Article Snippet: TOPFlash-mCherry reporter was optimized from TOPFlash-GFP reporter (Addgene, #35489).

Techniques: Expressing, Reporter Assay, Recombinant, Incubation, Concentration Assay, Activation Assay, Flow Cytometry, Agarose Gel Electrophoresis, Derivative Assay, Fluorescence

Journal: Cancer cell

Article Title: Synthetic Lethality of Wnt Pathway Activation and Asparaginase in Drug-Resistant Acute Leukemias

doi: 10.1016/j.ccell.2019.03.004

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: The 7TGC (7xTcf-eGFP//SV40-mCherry) TOPFlash reporter was a gift from Roel Nusse ( https://www.Addgene.org/24304/ ).

Techniques: Virus, Recombinant, Membrane, Staining, Caspase-Glo Assay, Purification, Cell Culture, Mutagenesis, Expressing, Plasmid Preparation, Software

Journal: Cancer cell

Article Title: Synthetic Lethality of Wnt Pathway Activation and Asparaginase in Drug-Resistant Acute Leukemias

doi: 10.1016/j.ccell.2019.03.004

Figure Lengend Snippet: (A) CCRF-CEM cells transduced with a constitutively active β-catenin were analyzed for expression of the indicated proteins by Western blot analysis (left), activity of a TOPFlash-EGFP reporter of β-catenin dependent transcriptional activity (mid), and transduction efficiency was assessed by immunostaining for β-catenin (right). Statistical significance was calculated using a two-sided Welch t-test. * p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; **** p ≤ 0.0001. n.s., p > 0.05.

Article Snippet: The 7TGC (7xTcf-eGFP//SV40-mCherry) TOPFlash reporter was a gift from Roel Nusse ( https://www.Addgene.org/24304/ ).

Techniques: Transduction, Expressing, Western Blot, Activity Assay, Immunostaining

Journal: Cancer cell

Article Title: Synthetic Lethality of Wnt Pathway Activation and Asparaginase in Drug-Resistant Acute Leukemias

doi: 10.1016/j.ccell.2019.03.004

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: The 7TGC (7xTcf-eGFP//SV40-mCherry) TOPFlash reporter was a gift from Roel Nusse ( https://www.Addgene.org/24304/ ).

Techniques: Recombinant, Staining, Caspase-Glo Assay, Purification, Cell Culture, Mutagenesis, Expressing, Plasmid Preparation, Software