ABSTRACT
Radioimmunoassay is a general method by which the concentration of virtually any substance can be determined. The principle on which it is based is summarized in the competing reactions shown in Figure
Ag — Ab Ag Ag*
Ag* • AbAb
17.1. The concentration of unlabeled antigen in the unknown sample is obtained by comparing its inhibitory effect on the binding of labeled antigen to a limited amount of antibody with the inhibitory effect of known standards. A typical RIA is performed by the simultaneous preparation of standard and unknown mixtures in test tubes. To these tubes are added a fixed amount of labeled antigen and a fixed amount of antiserum. After an appropriate reaction time, the antibody-bound (B) and free (F) fractions of the labeled antigen are separated by one of many different techniques. The B/F ratios in the standards are plotted as a function of the concentration of unlabeled antigen (“standard curve”), and the concentration of antigen in the unknown sample is determined by comparing the observed B/F ratio with the standard curve (Figure 17.2). Radioactive isotopes most frequently used for labeling are
H,
C,
S,
Co,
Se,
I, and
I (Table 17.1). Of these,
I offers useful characteristics for labeling and is very widely used. The RIA principle is not limited to immune systems, but can be extended to systems in which in place
of the specific antibody there is a specific reactor (that is, a binding substance) that might be, for instance, a specific binding protein in plasma,
an autoantibody,
an enzyme,
or a tissue receptor site.
