FIG. 4.

Immunocytochemistry of primary astrocytes infected with JCV reveals alterations in JCV protein levels upon JCV T-antigen and agnoprotein siRNA treatment. Cells were infected with JCV and treated at days 1, 5, and 10 postinfection with JCV T-antigen siRNA, JCV agnoprotein siRNA, or both. The cells were also treated with nonspecific agnoprotein siRNA. The cells were subcultured and plated onto poly-l-lysine-coated chamber slides (Falcon) on day 13 and were fixed on day 15 postinfection with ice-cold acetone for 3 min. Viral proteins were detected by immunocytochemistry as described previously (20), using the same primary antibodies as for the Western blot analysis (see the legend to Fig. 1). The proteins were visualized with fluorescein-conjugated secondary antibodies.