Hydroxyproline colorimetric assay kit

The total protein concentration in BALF was measured by a dye-binding assay using a commercial kit from ThermoFisher Scientific Incorporation (Waltham, MA, USA). The levels of TGFβ1, IL-1β, osteopontin, human MMP-2 (R&D Systems, Minneapolis, MN, USA), MCP-1, IL-13, interferon (IFN)-γ, IL-6 (BD Bioscience, BD opt-EIA kits, San Diego, CA, USA), MCP-3 (Preprotec, Rocky Hill, NJ, USA), connective tissue growth factor (CTGF) (Abgent, San Diego, CA, USA), Bax (LSBio, Seattle, WA, USA), and Bcl-2 (RayBiotech Life, inc, Peachtree Corners, GA, USA) were determined using commercial immunoassay kits according to the manufacturer’s instructions. Collagen type I was determined by enzyme immunoassay using an anti-collagen type I antibody and an anti-collagen type I biotin-conjugated antibody from Rockland Immunochemicals Incorporation (Limerick, PA, USA) following the manufacturer’s instructions. The level of total PDGF was measured by immunoassay as previously described [103 (link)]. The lung hydroxyproline content was determined by a colorimetric method using a commercial kit (Hydroxyproline colorimetric assay kit, BioVision, San Francisco, CA, USA) according to the manufacturer’s instructions.

Collagen deposition was measured by determination of hydroxyproline in the liver using the hydroxyproline colorimetric assay kit (Biovision, Milpitas, CA, USA). Briefly, liver samples were homogenized in distilled H₂O and the homogenates were hydrolysed in concentrated HCl by incubation at 120 °C for 3 h. Hydroxyproline contents in the dried hydrolysates were biochemically assessed according to the manufacturer's instructions.