Cdh1 610181

Immunohistochemistry was performed on formalin-fixed, paraffin-embedded human fetal lungs (T21 and non-T21) as previously described [27 (link)]. In brief, paraffin-embedded lungs were sectioned at 5 μm, rehydrated, and then boiled in a sodium citrate antigen retrieval solution for 12 min. A 15-min step in 3% H₂O₂ was performed to quench endogenous peroxidases to decrease background. Sections were incubated with the following primary antibodies overnight at 4 °C: CD31 (RB-10333-P0, 1:200; Neomarkers, Fremont, CA, USA), CDH1 (610181, 1:200; BD Biosciences, San José, CA, USA), Ki67 (RM-9106-S1, 1:200; Thermo Fisher Scientific, Fremont, CA, USA), LYVE1 (ab14917, 1:100; Abcam, Cambridge, UK), SOX2 (sc-17320, 1:100; Santa Cruz, Dallas, TX, USA), and SOX9 (AB5535, 1:500; Millipore, Burlington, MA, USA). Sections were then stained with appropriate fluorochrome-conjugated secondary antibodies in conjunction with Cy3-ACTA2 conjugated antibody (C6198, 1:200; Sigma-Aldrich, St Louis, MO, USA) when necessary. Slides were counterstained with DAPI (DE571; Life Technologies, Grand Island, NY, USA) and mounted using ProLong Diamond Antifade Mountant (Life Technologies).