A method for isolation of single-cell suspensions from lung in mice

Objective:To set up an effective method for isolation of single-cell suspensions from lung in mice.Methods:Isolation of lung cells was performed by type Ⅴ collagenase digestion.Briefly,lungs were finely dissected into small pieces using scissors and then digested with type Ⅴ collagenase(175 U·ml-1)for 1 h at 37℃.Then,single-cell suspensions were detected by cell counts and flow cytometry.Results:(1) By type Ⅴ collagenase digestion,higher single cell output rate,more intact cells,little cell fragments and little cell conglomeration were found in lung cell suspension.(2) It can be extracted(1-3)× 106 cells from 30 mg lung tissue in mice,and the percentage of living cells in lung single-cell suspension was 95% or more.(3) Flow cytometry showed a large number of lung single cells.Conclusion:The enzymatic digestion method by type Ⅴ collagenase is a simple,effective,stable and reliable method for the preparation of lung single-cell suspensions,which could be widely used to isolate lung tissues for in vitro functional tests.