OBJECTIVE: A bag-in-box system using a closed-circuit helium-dilution technique was used to determine the functional residual capacity during conventional mechanical ventilation. The purpose of this study was to determine whether the bag-in-box system could be used to measure mean lung volume without interrupting high-frequency oscillatory ventilation. DESIGN: Laboratory study. SETTING: Hospital intensive care medical research laboratory. SUBJECTS: Six mechanical lung models and ten New Zealand adult rabbits. INTERVENTIONS: The bag-in-box system was introduced into the respiratory circuit during the study. The rabbits were randomly divided into a healthy group and an acute lung injury group. Acute lung injury was induced by intravenously infusing oleic acid during conventional mechanical ventilation. MEASUREMENTS AND MAIN RESULTS: In a lung model study, a series of rebreathing processes were performed. By measuring the helium concentration in each process, the rebreathing time needed for equilibration was found to be about 50 secs. We compared the measured volume with a known lung volume to test the method's accuracy. The volumes differed by -0.5% to 4.4%. Measured and known volumes of different size lung models correlated well (p <.001). Equilibration of helium in the system occurred after 40-50 secs of rebreathing in healthy rabbits and 30-40 secs of rebreathing in rabbits with acute lung injury. Of 45 mean lung volume determinations in ten rabbits, the coefficient of variation ranged from 0.5% to 6.5%. When mean airway pressure changed, the mean lung volume alteration was detected by the method, but it was inconsistent in each mean airway pressure variation. CONCLUSION: A bag-in-box apparatus using a helium-dilution technique could be evaluated as a method to measure gas-containing lung volumes ranging from 10 to 60 mL in patients receiving high-frequency oscillatory ventilation.