In vivo localization of SP-Cpos/CCSPposcells. Frozen lung sections from ten 0 to 3 month old lambs were analyzed by immunohistochemistry for the expression of SP-C (green), CCSP (red) and the co-expression of SP-C and CCSP. The nuclei were stained with DAPI (blue). (A) Lung cross sections showed the expression of CCSP in the bronchiole (b) and SP-C in the alveoli (a) (100x magnification). (B) Immunostaining of rare SP-Cpos/CCSPpos cells in situ (400x magnification). Inset: enlargement of a SP-C/CCSP double-positive cell. (C) SP-C and CCSP expression from a representative lamb (# 1507) in ex vivo dissociated cells by flow cytometry showing the presence of a SP-Cpos/CCSPpos population. (D) Frequency of SP-Cpos, CCSPpos and SP-Cpos/CCSPpos cells from ten 0 to 3 month old lambs. The data are expressed as the mean (± SD) percentage of the cells expressing the cellular markers.