This report describes procedures to culture 1- to 2-mm-thick cross sections of lung lobes for periods of 4 to 6 weeks. Normal morphologic and macromolecular composition are maintained. Previous attempts to maintain adult peripheral lung cultures for periods beyond 7–10 days or to examine respiratory disorders in vitro other than acute changes have been generally unsuccessful. Eight different, supplemented, serum-free media, mixed with heated liquid agarose were infused into the airways of hamster and rat lungs. Cross sections were explanted onto squares of porous surgical packing material, placed in medium, and incubated for 4 to 6 weeks. The ability of each medium to maintain normal lung was assessed microscopically by quantitative image analysis and by biochemical analyses. The optimal medium formulation for each species is described. The adult peripheral lung culture system may provide toxicologists with a unique model for mechanistic and safety evaluations of potential lung toxicants.