Various factors influencing the apparent protein binding of quinidine were examined. Different binding values in rabbit plasma were obtained by equilibrium dialysis techniques employing three commonly used buffers. Binding values comparable to those found by ultrafiltration were achieved after dialysis against isotonic phosphate buffer for approximately 4 hr. Dialysis beyond 8 hr gave an increased free fraction with time. The reported effect of in vitro added heparin on plasma protein binding could be prevented by reducing the final concentration in blood from 20 to 5 U/ml, a concentration still sufficient to prevent clotting of the blood sample. Daily freezing and thawing of plasma samples over 1 week did not alter the binding of quinidine. The samples were stable for at least 2 months at -20 degrees.