RADIOIMMUNOASSAY OF 5-HYDROXYINDOLE ACETIC-ACID USING AN IODINATED DERIVATIVE

  • 1 January 1981
    • journal article
    • research article
    • Vol. 217  (3) , 791-797
Abstract
A radioimmunoassay for the main catabolite of serotonin, 5-hydroxyIAA (5-HIAA), was developed by using specific antibodies and iodinated derivative. The synthesis of a 125I-iodinated analog was performed by coupling 5-HIAA to [125I]glycyl-tyrosine without any contact between 5-HIAA and iodine or chloramine T. It was purified on a G25 Sephadex column and diluted in citrate buffer up to 2.5 .times. 105 cpm/ml. Antibodies were obtained by coupling 5-HIAA to human serum albumin with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide and tested by equilibrium dialysis. After the 3rd immunogen injection, the 4 rabbits gave antisera capable of binding 50% of iodinated 5-HIAA-glycyl-tyrosine at 1/2000 final dilution. A chemical conversion of the biological samples gives to the antigen molecules a better resemblance to the immunogen, thus confering a 100-fold gain in specificity and sensitivity. This assay allows 5-HIAA to be determined in small amounts of tissue, blood, CSF or perfusate without purification with a sensitivity threshold below 0.1 ng. Some applications in cat and rat are presented.