Computational protocol: Megakaryoblastic leukemia-1 is required for the development of bleomycin-induced pulmonary fibrosis

Similar protocols

Protocol publication

[…] Right lungs were inflated using formaldehyde and fixed for 24–48 hours before being moved to 70% ethanol. The lungs were sectioned and subjected to Masson’s trichrome staining and immunostaining for smooth muscle α-actin (α-SMA) or fibronectin. Secondary antibodies conjugated to either HRP (immunohistochemical images) or Alexafluor-488 (for quantitation of α-SMA) were utilized. The modified Ashcroft scale was used to quantify fibrosis, as previously described []. Quantitation of α-SMA immunostaining was performed by taking 10 images at 20X magnification per sample, using identical exposure times and thresholds on an Olympus 1X71 fluorescent microscope and Q imaging Retiga 2000R camera. ImageJ was used to measure integrated density of the fluorescent signal []. Quantitation of fibronectin histology was performed by taking a 2X magnification image of each sample, using identical exposure times and thresholds on an Olympus SZH10 microscope and Olympus DP70 camera. ImageJ Immunohistochemistry Toolbox plugin was utilized to isolate the brown stain []. The resulting image was inverted, the lung was outlined and integrated density measured. Integrated density measurement was divided by the total area of the lung in order to account for differences in lung sizes. […]

Pipeline specifications

Software tools ImageJ, IHC Toolbox
Application Microscopic phenotype analysis
Organisms Mus musculus
Diseases Leukemia, Megakaryoblastic, Acute, Pulmonary Fibrosis, Wounds and Injuries, Idiopathic Pulmonary Fibrosis
Chemicals Bleomycin