Isolation and Primary Culture of Murine Alveolar Type II Cells

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

Research output: Contribution to journalArticle

252 Citations (Scopus)

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
StatePublished - 1996
Externally publishedYes

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Suspensions
Osmium Tetroxide
Pulmonary diseases
Alveolar Epithelial Cells
Streptavidin
Tannins
Endothelial cells
Keratins
Fibronectins
Cell culture
Sepharose
Rats
Contamination
Recovery
Antibodies
Staining and Labeling
Lung
Inbred C57BL Mouse
Lung Diseases
dispase

ASJC Scopus subject areas

  • Cell Biology
  • Molecular Biology
  • Pulmonary and Respiratory Medicine

Cite this

Isolation and Primary Culture of Murine Alveolar Type II Cells. / Corti, Miriam; Brody, Arnold R.; Harrison, James H.

In: American Journal of Respiratory Cell and Molecular Biology, Vol. 14, No. 4, 1996, p. 309-315.

Research output: Contribution to journalArticle

Corti, Miriam ; Brody, Arnold R. ; Harrison, James H. / Isolation and Primary Culture of Murine Alveolar Type II Cells. In: American Journal of Respiratory Cell and Molecular Biology. 1996 ; Vol. 14, No. 4. pp. 309-315.
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