Western blot shows lysates of MCF-7 human breast cancer cell line untreated (-) or treated (+) with 100 ng/mL Recombinant Human IGF-I (Catalog # 291‑G1) for 20 minutes. PVDF membrane was probed with 0.5 µg/mL Human Phospho-p27/Kip1 (T157) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1555) followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). A specific band for Phospho-p27/Kip1 (T157) was detected at approximately 27 kDa (as indicated). The phospho-specificity of this antibody was supported by decreased labeling following treatment with 600 U lambda -phosphatase ( lambda -PPase) for 1 hour. This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.