Radioimmunoassays for serotonin and 5-hydroxyindole acetic acid

Radioimmunoassays for serotonin and 5-hydroxyindole acetic acid were developed. 1. High titer antibodies, having a well-defined high specificity, have been raised by coupling the side-chain of both molecules to human serum albumin. Serotonin is first converted into N-hemisuccinate, and then treated like 5-HIAA, namely, conjugated with HSA for the immunogen. 2. Synthesis of [125I] iodinated analogues was performed by coupling 5-HIAA or N-succinyl serotonin to glycyltyrosine without any contact between both molecules and the oxidizing reagents. 3. Chemical conversions of biological samples (by succinylation for 5-HT and amidation for 5-HIAA) were carried out. This critical step makes the antigen molecules resemble the immunogen more closely, thus allowing an appreciable gain in specificity and sensitivity. 4. These assays allow the rapid determination of 5-HT and 5-HIAA in small amounts of tissue, blood, cerebral spinal fluid or perfusate without any purification, with a sensitivity threshold of 50 pg.