Melatonin-like Immunoreactivity in the Presence of Different Chemicals as Determined by the Radioimmunoassay

To determine the nature of the molecule(s) that is responsible for the melatonin like immunoreactivity (MLI), we measured the effect of pretreatment of plasma samples with detergents, reducing agent, and proteinase K. Nonidet P-40 Triton X-100, and ethylacetate extraction had no effect, while sodium deoxycholate, sodium dodecyl sulfate, β-mercaptoethanol, ether extraction, proteinase K, and temperature increased the MLI. Since the radioimmunoassay (RIA) was sensitive to proteinase K, ionic detergents, and a reducing compound, we hypothesize that a proteinaceous molecule might be responsible for this MLI. We compared our column procedure for RIA of plasma melatonin (1) with procedures involving extraction with either ethylacetate or ether. In our hands preextraction of samples with organic solvents caused a loss of immunoreactivity. We also found that passing samples through the column is more efficient in eliminating interference in the melatonin assay than extracting samples either with ethylacetate or ether.