Chlamydia trachomatis grown in the yolk sac of embryonated eggs was purified by centrifugation on continuous isopycnic Renografin density gradients. A band of chlamydial particles with a buoyant density of 1.20 contained 70% of the starting particles, and electron microscopy revealed the virtual absence of contaminating egg material. Centrifugation on Renografin gradients caused only a moderate decrease in infectivity. For large-scale purification, infected yolk sac was centrifuged through Renografin solutions, resulting in greater than 60% recovery of starting chlamydial particles, but less than 1% recovery of the dry weight and protein.