Frontal sections of paraffinized knees joints were used for immunohistochemical analysis. Sections were deparaffinized and re-hydrated as previously described (24) . Primary antibodies used for immunolabelling are as follows: goat polyclonal Sox9 (AF-3075, R&D Systems), rabbit monoclonal GSK3alpha (ab40870, Abcam), rabbit polyclonal GSK3beta (12456S, Cell Signaling Technologies), mouse monoclonal type II collagen (Col2; 10R-C135b, Fitzgerald), rabbit polyclonal osteocalcin (ab93876, Abcam), rabbit polyclonal beta-catenin (bs-1165R, Bioss Antibodies) and rabbit polyclonal GLI1 (PA5-72942, Invitrogen, Thermo Fisher). Frontal sections without primary antibody were used as controls. Sections were incubated with was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission.
The copyright holder for this preprint (which this version posted April 7, 2020. ; https://doi.org/10.1101/2020.04.04.025700 doi: bioRxiv preprint respective HRP-conjugated anti-goat, anti-mouse and anti-rabbit IgG, and labelled proteins were probed with diaminobenzidine substrate (K3468, Dako) to reveal positive staining, followed by counterstaining in Methyl Green.
The copyright holder for this preprint (which this version posted April 7, 2020. ; https://doi.org/10.1101/2020.04.04.025700 doi: bioRxiv preprint respective HRP-conjugated anti-goat, anti-mouse and anti-rabbit IgG, and labelled proteins were probed with diaminobenzidine substrate (K3468, Dako) to reveal positive staining, followed by counterstaining in Methyl Green.