Fig 1: Dystrophin expression in cultured DMD myotubes by AO85 transfection. AO85 was transfected into the myotubes prepared as described in Figure 4 at a concentration of 50 nM. Seven days after the transfection, treated myotubes were harvested. After fixation with cold methanol, the fixed myotubes were incubated with polyclonal anti-dystrophin antibody (targeted to the C-terminal region; ab15277, Abcam, Cambridge, UK) and anti-myosin heavy chain (MHC) antibody (sc 101334, Santa Cruz Biotechnology, Dallas, TX, USA). The secondary antibodies were conjugated with Alexa Fluor 488 (Abcam) for dystrophin and Alexa Fluor 568 (Abcam) for the MHC. Immunohistochemical staining of dystrophin and MHC are shown. Before treatment, dystrophin was negative. After the transfection of AO85 (50 nM), myotubes were dystrophin positive, as was MHC staining. A quantitative assessment of dystrophin expression was not performed. Studies on DMD patients in this article were done after obtaining informed consent, in addition to approval from the local ethics committee.