Chromatin Immunoprecipitations (ChIP) were performed with cross-linked chromatin samples from 4 X 10⁶ of HT1080 cells treated with IFNγ for 30 minutes with either A)1:50 dilution of Go-ChIP-Grade™ Purified anti-STAT1 Phospho (Tyr701) (Clone A17012A) or B) equal amount of Purified Mouse IgG1, κ Isotype Control Antibody (Clone MG1-45, Cat. No. 401401) by using Go-ChIP-Grade™ Protein G Enzymatic Kit (Cat. No. 699904). The enriched DNA was purified and quantified by real-time qPCR using primers targeting human TAP1 gene region and α-Satellite repeats. The amount of immunoprecipitated DNA in each sample is represented as signal relative to total amount of input chromatin.

Total cell lysates (15 µg protein) from serum-starved HeLa S3 cells treated without (Lane 1) or with (Lane 2) 10 ng/mL human IFN-α2 (Cat. No. 592702) for 10 minutes were resolved by 8% Bis-Tris gel electrophoresis, transferred to nitrocellulose, and probed with 2 µg/mL (1:250 dilution) purified anti-STAT1 Phospho (Tyr701) antibody (clone A17012A). Proteins were visualized by chemiluminescence detection using HRP goat anti-mouse-IgG (Cat. No. 405301) at a 1:3000 dilution. Equal STAT1 loading was confirmed using a pan anti-STAT1 antibody that recognizes both STAT1 α (87 kD) and β (83 kD) isoforms.

Total cell lysates (15 µg protein) from serum-starved HeLa S3 cells treated without (Lane 1) or with (Lane 2) 10 ng/mL human IFN-α2 (Cat. No. 592702) for 10 minutes were resolved by 8% Bis-Tris gel electrophoresis, transferred to nitrocellulose, and probed with the indicated concentration of purified anti-STAT1 Phospho (Tyr701) antibody (clone A17012A). Proteins were visualized by chemiluminescence detection using HRP goat anti-mouse-IgG (Cat. No. 405301) at a 1:3000 dilution. Equal protein loading was confirmed using Direct-Blot™ HRP anti-β-Actin antibody (Cat. No. 643807) used at a 1:5000 dilution.

Human peripheral blood mononuclear cells were treated with (top), or without (bottom) Recombinant Human Interferon-γ (Cat No. 570204) for 15 minutes, fixed with Fixation Buffer (Cat. No. 420801), permeabilized with True-Phos™ Perm Buffer (Cat. No. 425401) then stained with CD4 (clone A161A1) APC and purified anti-STAT1 Phospho (Tyr701) (clone A17012A), followed by anti-mouse IgG PE. Data was gated on lymphocyte and monocyte populations.