This study evaluates the alveolar stereological profile of immature rabbit lungs after treatment with various surfactant preparations, deliberately modified to represent different rates of film adsorption, minimum surface tension and compressibility. Surfactant was isolated from porcine or bovine lungs, and some of these preparations were enriched with dipalmitoylphosphatidylcholine and other synthetic lipids. Alveolar stereological parameters were evaluated in histological lung sections by computerized interactive image analysis. Surfactant treatment enhanced lung expansion, with a significant correlation coefficient between dynamic compliance and alveolar volume density in all groups of animals. Surfactant treatment also significantly increased alveolar image area. Enrichment of surfactant with synthetic lipids did not improve stereological parameters, but caused a significant increase in the coefficient of variation for alveolar perimeter and image area, suggesting more heterogeneous expansion pattern. Experimental evaluation of exogenous surfactants in immature newborn animals should include assessment of alveolar stereological parameters to detect deviations from the uniform expansion pattern seen after treatment with an optimal surfactant substitute.