Image cytometry-based quantification protocol of human pulmonary arterial endothelial cells in lab-fabricated multichannel microfluidic devices.

Here, we present a protocol for automated quantification and viability analysis of pulmonary arterial cells (PACs) using a cellular image cytometer. We describe steps for obtaining PACs from the Pulmonary Hypertension Breakthrough Initiative, culturing and seeding them on microfluidic chips, and integrating chips into 6-well plates. Cells are stained with Hoechst 33342 and propidium iodide for live/dead analysis. Automated imaging and software-assisted quantification standardize workflows and broaden applications in pulmonary vascular research. For complete details on the use and execution of this protocol, please refer to Al Hilal et al.1.