Tendon Histology and Immunohistochemistry

Tendon tissues were fixed in 4% paraformaldehyde, dehydrated, and embedded in paraffin. Serial coronal sections (5 μm thick) were prepared from intrasynovial flexor tendons at 14 days after repair and intact intrasynovial and extrasynovial flexor tendons as previously described (N=2 per group).^{11 (link)} Repaired tendons were subjected to pentachrome staining^{13 (link)} (American MasterTech, Lodi, CA) for overall healing response including collagen deposition. Intact intrasynovial and extrasynovial tendons were subjected to CD146 immunohistochemistry as previously described.^{11 (link)} In brief, after heat-induced antigen retrieval, sections were incubated with rabbit anti-CD146 antibodies (Abcam, Cambridge, UK; #Ab75769, 1:200 dilution) at 4°C overnight. After three washes with 0.05% Triton X-100 in PBS (PBST), the sections were further incubated with biotinylated anti-rabbit IgG (Sigma Aldrich; 1:400 dilution) at room temperature for 1 hour, followed by ExtrAvidin-Peroxidase (Sigma Aldrich; 1:100 dilution) for 30 min. After three washes with PBST, the sections were developed with a DAB-Plus Substrate Kit (Life Technologies, Carlsbad, CA) and then counterstained with Mayer’s hematoxylin.