Visualizing Rotavirus and β-catenin in Infected Cells

Caco-2 and MA104 cells were cultured on glass coverslips and incubated after infection with RV-SA11 for 6 and 12 h. These cells were then fixed with 4% w/v paraformaldehyde in PBS and processed for the immunofluorescence technique, as mentioned previously [8 (link)]. Cells were incubated with the antibody dilutions specific to RV-VP6 and β-catenin (1:100) at 4 °C overnight, followed by incubation with FITC- (green) and Rhodamine- (red) conjugated secondary antibodies, respectively, in the dark for 2 h in a 37 °C incubator. The nuclei in the cells were stained with Vectashield containing 4′,6′-diamidino-2-phenylindole (DAPI-blue) and visualized with the help of the Axioplan confocal microscope (Carl Zeiss, Jena, Germany). The ZEN Blue software v3.1 (Carl Zeiss Microscopy, Jena, Germany) was used to process the microscopic images stored in the RGB format. In order to compare different images appropriately, these images were captured under identical excitation and detection settings in one session.