Analysis of pulmonary structure-function relationships by microscopy requires that the lung be fixed under carefully controlled physiological conditions, since lung structure is extremely responsive to the relationship between airway and vascular pressures. Unfortunately, standard post-mortem fixation techniques leave some doubt as to the exact relationship between these pressures during fixation. This problem can be circumvented by stabilizing lung structure by rapid freezing under carefully controlled physiologic conditions. Using ethylene glycol in a freeze substitution technique we have developed procedures which yield a degree of preservation compatible with the high degree of resolution of the electron microscope. These can be used to obtain a more detailed understanding of pulmonary structure-function relationships under well-defined physiological conditions.