The purpose of this study was to develop an optimal technique for performing sequential bronchoalveolar lavage (BAL) in a murine animal model. Two general anesthetic regimens and four operative techniques of BAL were tested. Anesthesia by intraperitoneal injection of ketamine hydrochloride (100 mg/kg body wt) resulted in death for four of ten subjects, whereas inhalation of diethyl ether led to death for one of ten subjects. BAL using a balloon catheter under bronchoscopic guidance was comparable with postmortal lavage, tolerated better, and resulted in superior cell retrieval with respect to cell differential (macrophages: 95 +/- 2.3; lymphocytes: 3 +/- 1.2; polymorphonuclear lymphocytes [PNL]: 1.2 +/- 1.4) compared with two other techniques using a bent metal tube/polyethylene tubing combination (macrophages: 19.3 +/- 27.4; lymphocytes: 3.8 +/- 4.3; PNL: 35.5 +/- 35.5) and a bronchoscope/polyethylene tubing combination (macrophages: 11.1 +/- 25.5; lymphocytes: 0.7 +/- 1.0; PNL: 55.8 +/- 41.0). The BAL fluid contained significantly more alveolar macrophages and fewer PNL and epithelial cells (p = 0.0001, p = 0.0025, p = 0.02, respectively). We conclude that the technique using a balloon catheter under bronchoscopic guidance during inhalation of diethyl ether is the procedure of choice and results in a representative sample of BAL.