The lipid layer of the tear film is associated with preventing evaporative loss. The ability of human Meibomian lipids to reduce evaporation in vitro was tested. Films of human or animal Meibomian lipids or mixtures of cholesterol and phosphatidylcholine were spread on the surface of either artificial buffer or on whole tears and placed on a mass balance that was enclosed in a sealed chamber. The temperature was adjusted to 37 degrees C and gas flow was controlled. Increasing the amounts of Meibomian lipids gave a very small reduction in evaporation. It was concluded from these in vitro experiments that prevention of evaporation from the tear film is not due to the Meibomian lipids alone, but is more likely to be due to a complex interaction between components of the aqueous and the Meibomian lipids.