Journal of Leukocyte BIology 44:391-401 (1988) © 1988 Alan R. Liss, Inc. Establishment and Characterization of Long-Term Cultured

Murine resident macrophages can proliferate in vitro when they are grown in coculture on a layer of mesothelial or endothelial type feeder cells. Resident macrophages were obtained from lung explants of C57B116 lpr/lpr mice and from spleen explants or perito-neal washing of Baib/c mice; the cells were seeded without further washing. After 3-4 weeks of culture, the macrophages began to proliferate on a confluent layer of feeder cells. The macrophages then could be collected in the fluid phase and reseeded for permanent culture after generation of a new feeder layer. These cells were characterized as macrophages by the following criteria: 1) their morphology, uitrastructure, and adher-ence properties; 2) more than 90 % of the macrophages phagocytized yeasts compared with less than 1 % of the feeder cells; 3) the presence of functional Fc and mannose receptors, nonspecific cytoplasmic esterases, and membrane ectoenzymes such as nicotinamide adenine dinucleotide (NAD) glycohydrolase and nucleotide pyrophospha-tase; 4) by cytofluorographic phenotype analysis with monoclonal antibodies, character-izing a normal macrophage population (MACi +, Fcrec +, H-2K +, THY1-, LYT2-, L3T4-). 5) by functional studies proving that the expanded macrophages could functio