Monoclonal rabbit anti alpha smooth muscle actin αsma

For histological observations, sections were stained with H&E. For immunohistochemistry, sections were deparaffinized followed by antigen retrieval via overnight incubation in 0.1 M Tris/HCl buffer, pH = 9.0 at 80°C.¹⁹ Sections were then washed with phosphate-buffered saline (PBS), permeabilized with 0.5% Triton X-100, and blocked with 5% Donkey serum (Jackson ImmunoResearch, Inc., West Grove, PA, USA) for 30 min at room temperature. After serum blocking, slides were incubated with 1:200 monoclonal rabbit anti-alpha smooth muscle actin (αSMA) (Abcam, Cambridge, MA, USA) in 1× PBS supplemented with 5% Donkey serum and 0.5% Triton X-100 at 4°C overnight. Detection of peripheral nerve axons was performed with incubation of 1:400 polyclonal rabbit anti-S100 (Dako North America, Inc., Carpinteria, CA, USA). Slides were then washed with PBS and incubated with 1:500 AlexaFluor^® 488 donkey anti-rabbit (Life Technologies, Carlsbad, CA, USA). Images were taken on an Olympus IX83 microscope at 20× with an Orca R2 camera (Hamamatsu Photonics K.K., Hamamatsu City, Japan) through Micro-Manager.^{20 (link)}