Phosphorylation of signal transduction molecules, Smad2 and Smad3, can be an important information for understanding of various biological functions of transforming growth factor (TGF)-β. TGF-β type I receptor and cyclin-dependent kinases phosphorylate both Smad2 and Smad3 at the C-terminals and at linker (middle) regions of them respectively (ref. 1). TGF-β signaling is mediated by Smad phosphoisoforms phophorylated at both the C-terminal and linker regions (ref. 2, 3). This monoclonal antibody against for Smad2L/3L (T220/T179) recognizes the part of phosphorylated threonin (Thr220 of Smad2 and Thr179 of Smad3) located in linker region of Smad2/3 specifically. It can be used for western blotting, immunoprecipitation and immunohistochemistry. It is applicable for enzyme biochemical analysis of TGF-β signaling and makes it possible to visualize the intermolecular reaction of phosphorylated Smad signaling in human tissues and monitor them in real-time. Thus, analysis of TGF-β signaling with this antibody is expected to be widely applied to cancer serearch (ref. 4) and fibrosis research (ref. 5), and to contribute to understanding the wide variety of life phenomina mediated by phosphoisoforms of Smad2/3.