Rate dialysis is used to obtain the free steroid fraction in undiluted plasma at 37 degrees C. The free steroid fraction is determined from the rate at which a small amount of tritiated steroid diffuses from plasma on one side of a semipermeable membrane into an identical plasma sample on the other side which lacks radioactive steroid. The method may be generally applicable to steroids since the cell permeability constant, which is a function of the volume of the dialysis cell and the area and diffusion properties of the membrane, was similar for seven steroids tested. The method requires only 0.3 ml of plasma, is simple and economical to perform, and enables up to 120 determinations to be made in one day. The free fractions of cortisol, progesterone, and estradiol-17 beta were measured in plasma pooled from pregnant and non-pregnant women and pregnant and lactating sows. The results were compared with those obtained for the same plasma pools by centrifugal ultrafiltration.