Morphometric estimates of diffusion capacity have traditionally been calculated for oxygen (DLO2) while physiologic techniques have been used to determine diffusion capacity for carbon monoxide (DLCO). Morphometric estimates of DLCO and Dm were determined for carbon monoxide so that the results can be directly compared to those obtained using physiologic techniques. Multiple gas rebreathing techniques were used to measure total lung diffusion capacity (DLCO), membrane component of diffusion capacity (Dm) and pulmonary capillary blood volume (Vc) in five anesthetized dogs. After the rebreathing measurements were made, the lungs were inflation fixed with glutaraldehyde and subjected to morphometric analysis. The morphometric estimate of diffusion capacity was three times higher than the physiologic estimate. The major components of diffusion capacity, Dm and theta Vc were found to be of approximately equal size when measured physiologically (Dm = 34 and theta Vc = 27 ml X min-1 X mm Hg-1). In contrast, the morphometric estimate of Dm was about six times higher than the morphometric estimate of theta Vc. The widely different ratios of the two major subcomponents of the total lung diffusion capacity as measured by these two different techniques suggest that the same lung function is not being accurately measured by at least one of these two techniques. Differences in results for DLCO cannot be explained by differences in the physiologic state of the animal at the time the measurement was made. Morphometric analysis predicts that Vc is the major term affecting the value of DLCO whereas the physiologic techniques predict that both Dm and Vc contribute substantially to the value of DLCO.