A study has been performed to investigate the tissue of origin of an endogenous plasma factor which has been previously shown to bind to an antidigoxin antibody raised in this laboratory. Adult male rats were used to provide tissue, which was homogenized ultrasonically and extracted on C18 minicolumns. The extracts were then assayed in a RIA for digoxin, and the ability of tissue extracts to displace radioiodinated digoxin from its antibody was determined. Consistent displacement was produced by extracts of whole adrenal gland, but not by whole brain and brain regions, anterior or posterior pituitary gland, skeletal muscle, aorta, testis, kidney, spleen, atrium, or ventricle. In some samples, extracts of liver also produced displacement of digoxin tracer. Further experiments were performed to determine whether bilateral adrenalectomy was able to influence plasma levels of the endogenous digitalis-like factor. Forty-eight hours after adrenalectomy, plasma levels of the factor were significantly reduced in whole plasma and ether extracts of plasma and were insignificantly lower in C18 extracts of plasma. These findings suggest that the endogenous factor we have studied previously in plasma may be derived from the adrenal gland.