Physiology has spawned many biological sciences, amongst them my own field of pharmacology. No man has made a more important contribution to the fields of physiology and pharmacology than Sir Henry Dale (1875( -1968 (4)), biological techniques and bioassay have contributed very substantially to the development of the field. Bioassay has provided crucial information on the role of the lungs in the removal of circulating prostaglandins (5), the participation of prostaglandins in inflammatory reactions (6, 7), the contribution of prostaglandins to the autoregulation and maintenance of blood flow to the kidney (8-10), the inhibitory effect of aspirin-like drugs on the biosynthesis of prostaglandins (11-13), the mediation of pyrogen fever by prostaglandins (14), and the release of rabbit aorta-contracting substance (RCS; now identified as thromboxane A2, TXA2) from lungs during anaphylaxis (15, 16). Moreover in 1976, bioassay made possible the discovery of PGX, now renamed prostacyclin (PGI2), the latest member of the prostaglandin family (17)(18)(19)(20). Indeed, it is doubtful whether the biological significance of any of the unstable products of arachidonic acid metabolism would have been recognized without bioassay techniques. With extraordinary simplicity and convenience, by its very nature, bioassay distinguishes between the important biologically active compounds and their closely related but biologically unimportant metabolites.In this review I shall discuss the development of the cascade superfusion bioassay technique and some of the discoveries and concepts which arose from its application, leading up to the discovery and development of prostacyclin. The effects of prostacyclin in man and its clinical assessment (another application of bioassay) will also be discussed.Cascade superfusion bioassay (a) Development Most uses of bioassay involving smooth muscle demand high sensitivity and specificity. These aspects have been achieved first, by limiting the volume of fluid bathing the isolated tissue and second, by using an assay organ sensitive to, and relatively specific for, the test substances under study. Further specificity can be achieved by using a combination of several tissues which present a characteristic pattern of response to the test substance or substances. This takes advantage of the principle of parallel pharmacological assay, regarded by Gaddum (21) as strong evidence for the identity of a compound.Magnus (22) introduced the idea of suspending an isolated portion of smooth muscle in a chamber