Calcium ion dependence of the 2-site immunoradiometric assay of Moore's S-100 protein

The two-site immunoradiometric assay (two-site IRMA) for a specific protein of the nervous system, S-100, was carried out by reaction of the S-100 protein solution with a solid-phase anti(S-100) followed by a second reaction in which the insoluble product was incubated with purified, radioactive anti(S-100). Unreacted labeled antibodies remained in solution and were washed away. As the amount of S-100 increased, the radioactivity in the solid phase increased. The most significant assay variable was the effect of calcium on the assay dose-response. 0.1 mM-EDTA caused a total inhibition of the dose-response curve which was reversed by increasing the concentration of calcium ions. The dose-response reached a maximum at 1.0 mM-Ca2+, then became progressively inhibited as the Ca2+ concentration was increased further. Previous immunochemical studies of S-100 which did not allow for this effect must now be interpreted with caution. (author)