Isolation and Primary Culture of Murine Alveolar Type II Cells

Miriam Corti, Arnold R. Brody, James H. Harrison

    Research output: Contribution to journalArticlepeer-review

    263 Scopus citations

    Abstract

    Previous attempts to culture mouse alveolar type II (ATII) cells have been hampered by limited purity and cell recovery. We have now obtained culturable ATII cells from female C57BL/6 mice at a purity of 92% ± 3 (mean ± SD; n = 20), with viabilities of 96% ± 2 and total yields of 5.1 ± 0.7 × 106 cells per mouse. Crude lung cell suspensions were prepared by intratracheal instillation of Dispase and agarose followed by mechanical disaggregation of the lungs. Crude cell suspensions were purified by negative selection using a biotinylated-antibody, streptavidin-coated biomagnetic particle system. Cell purities were determined by Pap staining and confirmed ultrastructurally. Purified ATII cells were cultured on fibronectin-coated chamber slides and maintained for up to 5 days in DMEM with 10% fetal bovine serum. Cultures exhibited minimal contamination by Clara cells, mesenchymal cells, or endothelial cells, and the epithelial nature of the cultures was confirmed by positive cytokeratin staining in at least 97% of the cells through day 5. Day 3 cultures demonstrated osmium tetroxide/tannic acid-stained granules consistent with lamellar bodies in 76% ± 3.6 of the cells. The cultures displayed features distinct from those previously described for adult rat ATII cells, including irregularly-shaped cells and the formation of numerous cytoplasmic projections in direct contact with other cells. These studies indicate that excellent yields of highly purified, culturable ATII cells can be obtained from genetically defined mice. These techniques may provide powerful new models for the study of parenchymal lung disease in vitro.

    Original languageEnglish (US)
    Pages (from-to)309-315
    Number of pages7
    JournalAmerican journal of respiratory cell and molecular biology
    Volume14
    Issue number4
    DOIs
    StatePublished - 1996

    ASJC Scopus subject areas

    • Molecular Biology
    • Pulmonary and Respiratory Medicine
    • Clinical Biochemistry
    • Cell Biology

    Fingerprint

    Dive into the research topics of 'Isolation and Primary Culture of Murine Alveolar Type II Cells'. Together they form a unique fingerprint.

    Cite this