Isolation of highly pure alveolar epithelial type I and type II cells from rat lungs.
There are no ideal cell lines available for alveolar epithelial type I and II cells (AEC I and II) at the present time. The current methods for isolating AEC I and II give limited purities. Here, we reported improved and reproducible methods for the isolation of highly pure AEC I and II from rat lungs. AEC I and II were released from lung tissues using different concentrations of elastase digestion. Macrophages and leukocytes were removed by rat IgG 'panning' and anti-rat leukocyte common antigen antibodies. For AEC II isolation, polyclonal rabbit anti-T1alpha (an AEC I apical membrane protein) antibodies were used to remove AEC I contamination. For AEC I isolation, positive immunomagnetic selection by polyclonal anti-T1alpha antibodies was used. The purities of AEC I and II were 91 +/- 4 and 97 +/- 1%, respectively. The yield per rat was approximately 2 x 10(6) for AEC I and approximately 33 x 10(6) for AEC II. The viabilities of these cell preparations were more than 96%. The protocol for AEC II isolation is also suitable to obtain pure AEC II (93-95%) from hyperoxia-injured and recovering lungs. The purified AEC I and II can be used for gene expression profiling and functional studies. It also offers an important tool to the field of lung biology.
Chen
Jiwang
J
Department of Physiological Sciences, Oklahoma State University, Stillwater, OK 74078, USA.
Chen
Zhongming
Z
Narasaraju
Telugu
T
Jin
Nili
N
Liu
Lin
L
D000818:Animals; D000906:Antibodies; D002469:Cell Separation / Q000379:methods*; D002470:Cell Survival; D004847:Epithelial Cells / Q000166:cytology / Q000502:physiology; D018496:Hyperoxia / Q000473:pathology / Q000503:physiopathology; D018189:Immunomagnetic Separation / Q000379:methods; D055370:Lung Injury; D008297:Male; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000276:immunology; D008854:Microscopy, Electron; D010196:Pancreatic Elastase; D011650:Pulmonary Alveoli / Q000166:cytology* / Q000502:physiology; D037661:Pulmonary Surfactant-Associated Proteins / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results; D000098853:Podoplanin
D000906:Antibodies; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D010196:Pancreatic Elastase; D037661:Pulmonary Surfactant-Associated Proteins; C092025:Pdpn protein, rat; D000098853:Podoplanin