Standardization of incubation time and temperature for eight radioimmunoassays

We wanted to know if incubation time and temperature for many radioimmunoassay methods could be standardized, to decrease assay time and so improve efficiency. Percentage binding was determined for triiodothyronine, thyroxine, digoxin, digitoxin, testosterone, aldosterone, estradiol, and diphenylhydantoin when the reaction mixtures were incubated at 6-8, 22, or 37 degrees C for various periods of time. In all methods tested, binding of antigen to antibody was greatest when the reaction mixtures were incubated at 6-8 degrees C, less at 22 degrees C, and least at 36 degrees C. Maximum binding occurred within 30-60 min at each temperature for all methods tested. Hence, it is possible to standardize the incubation time and temperature for these eight radioimmunoassay methods to 60 min at 6-8 degrees C.