Rabbit anti pdgfrα

Muscles were cut into 10 μm muscle cross-sections and mounted on glass slides to be stained, as described below. Trichrome staining was conducted by the Histology Core Facility at the University of Ottawa following Masson’s Trichrome protocol⁴⁸ . For immunofluorescence analyses, muscle cross-sections were fixed with 2% paraformaldehyde (PFA) for 7 minutes, permeabilized using 0.1% Triton X-100 (Thermo Fisher Scientific, Waltham, MA) for 10 minutes at room temperature, blocked using either the M.O.M Blocking kit (Pax7; Vector Labs, Burlingame, CA), DAKO Serum-Free Protein Block (PDGFRα; Agilent, Santa Clara, CA), or standard serum block (Laminin, Perilipin, and p-NFkB; PBS with 5% NGS, 1% BSA, and 0.1% Triton X-100). Sections were incubated with primary antibody overnight at 4 °C (mouse anti-Pax7, DSHB, neat; rabbit anti-MyoD, Abcam, 1:200; rat anti-Laminin, ThermoFisher Scientific, 1:200; rabbit anti-PDGFRα, R&D Systems; 1:250; rabbit anti-phospho-NFĸB, Cell Signaling, 1:200; Perilipin, Cell Signaling, 1:100). Appropriate Alexa secondary antibodies (Invitrogen, Carlsbad, CA) were used for detection of each primary antibody (1:300), while nuclei were counterstained using 4,6-diamidino-2-phenylindole (DAPI, 1:10000).

The following antibodies were used: Rat anti-BrdU (1:100, Abcam, ab6326); Rabbit anti-Ki67 (1:100, Abcam, ab16667); Rabbit anti-H3pS10 (1:150, Epitomics, 1173-1); Mouse anti-Setd4 (1:50, Santa Cruz Biotechnology, sc-514060); Rabbit anti-H4K20me3 (1:200, Abcam, ab9053); Mouse anti-HP1α (1:200, EMD Millipore Corp, 05-689); Rabbit anti-H3K9ac (1:200, Epitomics, 1328-1); Mouse anti-Troponin T (1:100, Invitrogen, MA5-12960); Mouse anti-sarcomerica-actin (1:100, Sigma, A2172); Rat anti-CD31 (1:200, BD Biosciences, 5553370); Goat anti-VE-cadherin (1:200, RD, AF1002); Rabbit anti-PDGFRα (1:100, RD, AF1062); Rabbit anti-Sox9 (1:100, Abcam, ab185230); Rabbit anti-α-SMA (1:150, Sigma, A2547); Rabbit anti-FABP4 (1:150, Abcam, ab92501); Rat anti-c-Kit (1:100, Abcam, ab25022).