Abstract
Isolation of relatively pure populations of alveolar type II epithelial cells has previously only been possible by density gradient centrifugation techniques which exploit the low buoyant density conferred on such cells by lung surfactant containing lamellar bodies. We have used a multiparameter laser flow cytometer (cell sorter) to obtain similar populations of lamellar body containing cells from single cell suspensions made from rabbit lungs subjected to enzymatic dissociation by trypsin and elastase. The lamellar body cells are sorted on the basis of two parameters: low angle light scatter, and green fluorescence from the lipid seeking probe phosphine-3R. Cells do not separate well on the sorter when either of these two parameters is used alone. For example,alveolar macrophages scatter light in an overlapping range with lamellar body cells, and various kinds of cell debris stain with phosphine-3R in the same intensity region as viable cells with lamellar bodies. However, when both light scatter and fluorescence are considered simultaneously, the multiparameter cell sorter gives a highly purified population of lamellar body cells in a regime of high light scatter and high fluorescence. Presently we sort at a rate slightly less than 107 lamellar body cells per hour, certainly less than obtainable with gradient separation methods. However, the multiparameter cell sorter has the advantage of near absolute purity of type II cell populations obtained, and has the potential to isolate developing lung cells with few lamellar bodies which would not readily segregate by buoyant density.
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Finkelstein, J., Leary, J., Notter, R. et al. 1654 ISOLATION OF ALVEOLAR TYPE II CELLS WITH A LASER FLOW CYTOMETER. Pediatr Res 15 (Suppl 4), 719 (1981). https://doi.org/10.1203/00006450-198104001-01672
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DOI: https://doi.org/10.1203/00006450-198104001-01672