Differences in subcellular localization between STAT1-WT and STAT1-KR after IFN-γ stimulation. U3A STAT1-WT (A) and STAT1-KR (B) stable clones were serum starved overnight and stimulated with IFN-γ (100 ng/mL) for the indicated time points. STAT1 subcellular localization was visualized by immunofluorescence staining using specific anti-STAT1 antibody (left column of each panel) and anti–phospho-STAT1 (Y701) antibody (right column). Images were visualized under an Olympus IX70 confocal microscope equipped with a 100×/1.4 oil immersion objective lens (Olympus, Melville, NY) and an ULTRAPix ICX 085 camera (PerkinElmer, Wellesley, MA). Green emission was detected through 488 mm/10 mm and 525 mm/50 mm filters; red emission, through 568 mm/10 mm and 608 mm/45 mm filters (all filters from Perkin-Elmer Life Sciences, Cambridge, United Kingdom). UltraView 4.0 software (Perkin-Elmer) was used for image acquisition, and PhotoEditor software for Microsoft XP Office (Microsoft, Redmond, WA) was used for image analysis.