Characterization of site-directed antisera against endothelin-converting enzymes

Site-directed antisera have been developed against the two endothelin-converting enzyme-1 (ECE-1) isoforms cloned to date in humans, ECE-1α and ECE-1β. Antisera were raised in rabbits against synthetic peptides corresponding to the deduced amino acid sequences that differ between ECE-1α and ECE-1β. Antisera were highly selective for their corresponding antigen (titer 1 x 10 4 ) and did not detect ET-I or big ET-I. Furthermore, no detectable crossreactivity was observed between the different site-specific antisera and the other immunizing peptides, suggesting that the antisera would be selective for ECE-1α and ECE-1β. Standard displacement curves have been developed to determine the levels of immunoreactive ECE-1α and ECE-1β in solubilized microsomal fractions of human tissue. In conclusion, we have described the first production and characterization of site-directed antisera raised against ECE-1α and ECE-1β capable of discriminating between the two ECE-1 isoforms. Furthermore, using these antisera, we have found that ECE-1α appears to be the predominant isoform in human tissue.