Grimpe, P., Melcher, L., Gröbe, L., Hühn, J., Boehme, J. D., Bruder, D
Summary
This protocol presents a flow cytometry-based method to isolate viable alveolar type II epithelial cells (AECII) from neonatal, juvenile, or adult murine lungs (infected or non-infected). This approach enables downstream molecular and/or functional studies of AECII to identify their roles during lung inflammation and infection.
Abstract
Alveolar type II epithelial cells (AECII) critically contribute to immune regulation in the lung and thus are key elements orchestrating pulmonary homeostasis. These functions render AECII particularly relevant in the context of infection with respiratory viruses, which target the lower airway epithelium and disrupt alveolar integrity. Preclinical mouse models are vital tools to elucidate the roles of AECII in the context of lung inflammation and/or infection. This protocol presents a detailed and reproducible pipeline for isolating viable and highly purified AECII from murine lungs at neonatal, juvenile, and adult stages. The methodology combines mechanical dissociation, enzymatic digestion, and subsequent fluorescence-activated cell sorting (FACS) to yield highly enriched AECII populations proven to be suitable for downstream molecular and functional analyses. For applications involving pathogenic viruses, the protocol includes an optional paraformaldehyde (PFA) fixation step, enabling safe handling under lower biosafety conditions without compromising RNA integrity. With optimized cell recovery across developmental stages, yielding up to 1.0 × 106 AECII cells per adult lung, this protocol supports preclinical studies of, e.g., viral pathogenesis, epithelial function, and regenerative capacity.
