Fig. 6.

Nuclear translocation of AhR. LS180 cells were seeded on chamber slides and cultured for 2 days. Then, the cells were incubated for 90 minutes with DMSO (0.1% v/v), TCDD (5 nM), 4-Me-indole (1, 10, and 100 μM), and 7-MeO-indole (1, 10, and 100 μM). Microscopic specimens were prepared according to the common protocol, using Alexa Fluor 488 labeled primary antibody against AhR, 4′,6-diamino-2-phenylindole, and VectaShield Antifade Mounting Medium. AhR was visualized and evaluated using a fluorescence microscope. Percentage of cells with nuclear AhR was calculated by visual comparison of antibody signal intensity in the nucleus and cytosol, when at least four random locations per sample with approximately 100 cells were evaluated. Experiments were performed in two consecutive cell passages, with all tested compounds in duplicate. The representative images are shown together with an inset containing the total and AhR-positive counts of cells.