Immunostaining of Tibialis Anterior Muscle Sections

Tibialis anterior (TA) muscle sections were prepared for staining essentially as previously described^{6 (link),48 (link)}. For OSMR staining, whole TA muscles were first fixed with 0.5% paraformaldehyde/PBS for 2 h at 4 C, followed by incubation in 20% sucrose/PBS overnight at 4 C. 10 μM sections were cut for all stainings. Antibodies were: anti-laminin (Millipore, catalog #05⁻206, 1:250), anti-GFP (Invitrogen, catalog #A11122, 1:200), anti-OSMR (R&D, catalog #AF665, 1:200), anti-Pax7 (Santa Cruz Biotechnology, catalog #sc81648, 1:50; or Developmental Studies Hybridoma Bank, 2 μg/mL final concentration), AlexaFluor 594–conjugated donkey anti-rat IgG1 and AlexaFluor 488–conjugated donkey anti-rabbit (Jackson ImmunoResearch, catalog # 712-585-150 and 711-545-152 respectively, 1:200 each). Nuclei were counterstained with either DAPI (Invitrogen) or TO-PRO-3 (Invitrogen). Images were acquired with an AxioPlan2 epifluorescent microscope (Carl Zeiss) with ORCA-ER digital camera (Hamamatsu Photonics).