Abstract Efficient mixing of inspired gas with the resident gas of the lung is an essential requirement of effective respiration. This review focuses on one method for quantifying ventilation inhomogeneity: the multiple-breath inert gas washout (MBW). MBW has been employed as a research tool in adults and school age children for more than 50 years. Modifications allowing data collection in infants and preschoolers have been described recently. Indices of overall ventilation inhomogeneity, such as the lung clearance index and moment ratios, are raised in many infants with lung disease of prematurity, and in young children with cystic fibrosis. These indices may be more sensitive than other lung function measures for the early detection of airway disease. We describe, for the first time, a development of the MBW analysis that allows calculation of acinar and conductive zone inhomogeneity indices in spontaneously breathing children. Although methodological and analytical issues remain, the future clinical and research applications of MBW justify accelerated research in this field.