Characterization of type II alveolar epithelial cells by flow cytometry and fluorescent markers

Type II alveolar epithelial cells play a crucial role in maintaining the structure and functions of pulmonary alveoli. A number of techniques have been described to isolate type II cells for in vitro studies; however, type II cell suspensions isolated by each technique are still contaminated by macrophages or monocytes. The present studies describe the use of flow cytometry to accurately characterize the composition of these cell suspensions. With freshly isolated type II cell suspensions, type II cells could be distinguished from macrophages and monocytes by two methods: (1) the combination of natural fluorescence and orthogonal light scatter, or (2) the use of monoclonal antibodies OX-1 (directed against a common leukocyte antigen present on rat macrophages and monocytes) and PKK-1 (directed against cytokeratin intermediate filaments present in type II cells). With cultured type II cells, the combination of natural fluorescence and orthogonal light scatter did not distinguish between type II cells and macrophages or monocytes; however, the monoclonal antibodies OX-I and PKK-1 continued to distinguish between these cell types. As an example of the use of these techniques, the methods were used to define the sequential expression of class I and II major histocompatibility antigens on both type II cells and on macrophages or monocytes in the same cell preparations. These methods are of potential value in isolating pure populations either of type II cells or of macrophages or monocytes by cell sorting and in accurately identifying the cells present in type II cell suspensions or cultures.