Quantifying TGF-β1 Expression in Tracheal Scars

In situ TGF-β1 expression in the tracheal scar was assessed by immunohistochemistry (IHC) of the tracheal tissue sections used for microscopy with a rabbit polyclonal anti-TGF-β1 antibody (Abcam ab25121, USA) using the biotin-streptavidin-peroxidase system (Vectastain Universal Quick Kit, Burlingame, CA). The sections were incubated with 3,3′-diaminobenzidine (DAB, BioCare Medical, USA) and counterstained with CAT hematoxylin (BioCare Medical, USA). After completing the staining, TGF-β1 expression was quantified in the entire sample circumference using the program ImageJ (http://rsbweb.nih.gov/ij/), which was developed by the National Institutes of Health (NIH), and the plugin IHC Profiler [17 (link)].