Cardiotrophin-1 (CT-1) is a new member of the interleukin (IL)-6 family of cytokines and one of the endogenous ligands for gp130 signaling pathways in the heart, which has potent hypertrophic and survival effects on cardiac myocytes. However, the clinical significance of CT-1 is poorly understood, mainly because there is no widely applicable specific and sensitive assay system for measuring plasma levels of circulating CT-1. We therefore developed a competitive radioimmunoassay (RIA) for human CT-1 with rabbit antiserum recognizing the N-terminus region of human CT-1 and using recombinant human CT-1 as a calibrator. The assay displays no cross-reactivities with any of the IL-6 family of cytokines including IL-11, leukemia inhibitory factor, ciliary neurotrophic factor, and oncostatin M. The lower detection limit in buffer was found to be 43 fmol/ml, and the working range was 120-8300 fmol/ml (CV < 15%). This RIA directly recognizes CT-1-like immunoreactivity in human plasma with a mean value of 571 75 fmol/ml (mean +/- SD) in healthy volunteers. The RIA coupled with gel filtration chromatographic analyses showed that the major molecular form of circulating CT-1 corresponds to recombinant full-length human CT-1. Moreover, there is a significant increase in the plasma CT-1 concentration from the aorta and coronary sinus, which clearly indicates that the heart secretes CT-1 via the coronary sinus into the peripheral circulation. This RIA should serve as a powerful tool for investigating the clinical significance of CT-1.