hPSC-AECs were incubated with collagenase IV (Sigma-Aldrich) for 2 hrs, followed
by treatment with cell dissociation buffer (Gibco, Grand Island, NY, USA) for 30
min at 37°C. Cells were passed through a 70 μm cell strainer and
incubated with following primary antibodies for 1 hr at 4°C: NKX2.1
(Abcam), carboxypeptidase M (CPM), epithelial cell adhesion molecule (EPCAM,
Santa Cruz, Dallas, TX, USA), Surfactant protein B (SFTPB, EMD Millipore,
Burlington, MA, USA), SFTPC (Abcam), T1α and aquaporin 5 (AQP5). Dead
cells were excluded by 7-aminoactinomycin D (BD Pharmingen, San Diego, CA, USA)
staining. Frequencies of AEC markers were measured using a FACSCantoTM II flow
cytometer (BD Bioscience), and acquired data were analyzed with FlowJo software
(Tree Star, Ashland, OR, USA).
by treatment with cell dissociation buffer (Gibco, Grand Island, NY, USA) for 30
min at 37°C. Cells were passed through a 70 μm cell strainer and
incubated with following primary antibodies for 1 hr at 4°C: NKX2.1
(Abcam), carboxypeptidase M (CPM), epithelial cell adhesion molecule (EPCAM,
Santa Cruz, Dallas, TX, USA), Surfactant protein B (SFTPB, EMD Millipore,
Burlington, MA, USA), SFTPC (Abcam), T1α and aquaporin 5 (AQP5). Dead
cells were excluded by 7-aminoactinomycin D (BD Pharmingen, San Diego, CA, USA)
staining. Frequencies of AEC markers were measured using a FACSCantoTM II flow
cytometer (BD Bioscience), and acquired data were analyzed with FlowJo software
(Tree Star, Ashland, OR, USA).