Protein extraction from formalin-fixed paraffin-embedded (FFPE) lung tissues and western blot analysis:
FFPE lung tissues (10 μm sections) from non-COVID-19 and COVID-19 patients were transferred into 1.5 ml centrifuge tubes, dewaxed in xylene for 10 min and centrifuged. T issue pellets were washed 3x with xylene and rehydrated with graded concentrations of ethanol (100 %, 85 % and 75 % for 2 min in each). After the ethanol wash, proteins in the tissue pellets were solubilized in Tris-SDS buffer, pH 8.0, by homogenization and sonication. Tissue homogenates were centrifuged at 10,000 g for 20 min and the supernatants were measured for total protein concentrations. Equal amounts of proteins were resolved by 12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis, followed by transfer onto nitrocellulose membrane. Membranes were then blocked with 5% bovine serum albumin and incubated with anti-elastin (1:1000 dilution) and anti-histone H3 Pan antibodies (loading control, 1:4000 dilution) overnight at 4 0C. Next day, membranes were washed in TBST and incubated with IRDye 800CW and IRDye 600CW conjugated secondary antibodies. The membranes were washed 3 times and signal was measured using the Odyssey Infrared Imaging System (LI-COR Biosciences).