Radioimmunoassay of melatonin in rat serum

• 1.

  1. Melatonin was coupled to albumin by means of a N_a-p-carboxybenzyl, a N_a-propionic acid, methylene, or a diazotized p-aminobenzoic acid bridge and injected into rabbits or sheep.

• 2.

  2. All coupling procedures stimulated antisera which bound melatonin with the greatest affinity; N-acetylserotonin and 6-hydroxymelatonin cross-reacted with the antisera at 3–5%. Other naturally occurring indolealkylamine derivatives showed less cross-reactivity.

• 3.

  3. Coupling the hapten melatonin to protein through a bridge at or near the indole N can stimulate antisera which are relatively specific to melatonin and suggest that antisera which bind specifically to any other individual indolealkylamine derivative could be produced using these coupling procedures.

• 4.

  4. Using the methylene bridged antisera, a radioimmunoassay for melatonin was developed and validated for rat serum. In addition to tests for reliability and parallelism, serum estimates of melatonin by radioimmunoassay correlated .98 with gas chromatography-mass spectrometry.

• 5.

  5. Serum melatonin levels have a characteristic 24-hour rhythm with a crest late in the dark period (L:D 12:12). Following the injection of 50 ug of melatonin, serum levels reach 5–20 fold higher concentrations than physiological levels.