The regional distribution of cholesterol, phospholipid and protein content was determined on pools of human lenses ranging from 13 to 68 years old. The study was undertaken to establish age matched controls for comparison with cataractous lenses. Future spectroscopic structure analysis of human lenses will be performed and the results related to chemical composition. The molar cholesterol to phospholipid ratio was 3.5 +/- 0.3 for human lens. This ratio is high for human tissue. The lens ratio increased from 2.2 +/- 0.3 in the equatorial region to 9.2 +/- 1.6 in the nuclear region. This trend was also observed in the bovine lens. The relative amount of protein increased concomitantly from 0.13 +/- 0.02 Kg protein per gram lipid in the equatorial region to 0.33 +/- 0.06 in the nucleus. The cholesterol to protein ratio remained constant throughout the lens at 0.073 +/- 0.003 Kg suggesting cholesterol could be associated with the crystallin proteins. In partially purified membrane preparations the cholesterol to phospholipid molar ratio was 2.6 +/- 0.2 and 3.2 +/- 0.2 for the cortex and nucleus respectively, three times lower than for the whole tissue. The high cholesterol content could account for the observed rigidity of membranes measured by infrared spectroscopic examination of the CH stretching band.