603129.fig.002a
(a)
603129.fig.002b
(b)
Figure 2: Results of the characterization of the vector construct SuperTop-eGFP4-1-5. (a) Microscopic analysis of HEKT-STE4-1-5 1C cells treated with either 0,1 mM, 1 mM, 10 mM LiCl (upper row, from left to right) or 20 ng/mL, 50 ng/mL, 100 ng/mL WNT3A (lower row, from left to right), respectively. Representative images were taken 24 hours after treatment. The fluorescence signal was highest with 10 mM LiCl due to Wnt pathway activation. Treatment with either concentration of WNT3A also showed minor increase in fluorescence as compared to LiCl activation. Magnification 200x. (b) Quantitative analysis of Wnt pathway activation in stably transfected HEKT-STE4-1-5 1C cells. Cells were treated either with LiCl or WNT3A protein and fluorescence signal was monitored right after, 24 hours and 48 hours after treatment. Bars are showing the fluorescence signal in comparison to the untreated control, which was normalized to 1 (left axis). Data were normalized to cell number per well. The lines show the percentage difference in fluorescence signal from 0 h to 24 hours and from 24 hours to 48 hours measurements (right axis). In LiCl-treated cells the highest fluorescence increase is displayed 24 hours after treatment and with a concentration of 1 mM LiCl. Highest change in fluorescence after addition of WNT3A was monitored within a concentration of 50 ng/mL WNT3A, although the signal is lower as compared to the signal obtained in cells treated with 20 ng/mL WNT3A which is comparative to the signal in 1 mM LiCl-treated cells.