Human pulmonary surfactant contains four groups of apoproteins, SP-A, B, C and D. We determined the concentration of SP-A in the tracheal aspirate of newborn infants by a two-site simultaneous immunoassay with monoclonal antibodies, and used this assay to assess changes in surfactant in various clinical situations. SP-A concentrations were standardized per milligram of albumin in the aspirate. The ratio of SP-A/albumin (micrograms/mg) in tracheal aspirates of 18 preterm infants with respiratory distress syndrome (RDS), in which samples were obtained within 12 hours of birth, was significantly lower (0.2 +/- 0.1 microgram/mg, mean +/- S.D.) compared to a group of 20 non-RDS preterm infants of similar gestational age (15.8 +/- 7.4 micrograms/mg) (p less than 0.05). None of the RDS infants had a SP-A/albumin ratio above 1 microgram/mg within 12 hours of birth, but the ratio exceeded 5 micrograms/mg in all samples from non-RDS infants. The SP-A/albumin ratio significantly increased, however, at 48 to 72 hours after birth in infants with RDS (15.7 +/- 9.5 micrograms/mg). During the recovery phase of RDS, no difference was evident in the SP-A/albumin ratio in babies treated with artificial surfactant compared to those not treated.