Non-specific binding of insulin in an equilibrium binding assay of insulin antibodies

Abstract

In liquid phase assays for insulin binding antibodies (IBA), total binding of insulin is composed of specific and non-specific binding (NSB). Sometimes NSB is determined in serum of healthy individuals and then subtracted from total binding of IBA positive serum to obtain specific binding. This method does not take into account that NSB might vary from plasma to plasma. This possibility was investigated by means of a computerised non-linear curve fitting routine for the evaluation of measurement results of an (equilibrium) binding assay for IBA, which yields estimates of NSB for each plasma individually. From each of 19 insulin treated diabetic patients, 4 blood samples, taken at different points in time, were available for IBA and NSB measurement. It was found that inter-patient variance of NSB exceeded within-patient variance ( p < 0.01) and, in a number of instances, within-patient variance was greater than experimental variance. Our results indicate that it is advisable to use methods for IBA evaluation that take these NSB variations into account.