Measurement of and Correction for Incubation Damage in Radioimmunoassay

The role of incubation damage in the radioimmunoassay of human plasma insulin, human plasma growth hormone and rat plasma growth hormone was studied. Damage was measured as the fraction of labelled material which migrated from the origin in wick paper chromatography of samples without antibody. The experiments demonstrated, that recovery of added hormone differed increasingly from 100% as incubation damage increased. Recoveries were found to be too high using the double antibody technique, and too low using chromatographic technique. However, satisfactory recoveries could be obtained in chromatographic assay after correction for the experimentally determined incubation damage using the general formula: % Bound=100×(B‐KD)/ (100‐KD) where B is the apparent antibody‐bound fraction, D the damage fraction, both expressed as percent of the total radioactivity. K is determined experimentally.