The Measurement of Serum Thyroglobulin in the Presence of Thyroglobulin Antibodies

Abstract

In a significant proportion of patients with thyroid disorders, thyroglobulin antibodies (TgAb) invalidate double-antibody radioimmunoassays (RIA) for thyroglobulin (Tg). A modified RIA procedure for determining serum Tg in the presence of TgAb, is described. Two measurements are made on each sample. a Ratio of immunoglobulin (Ig)-bound thyroglobulin to free thyroglobulin (bound:free ratio). This is measured by the distribution of a trace quantity of 125I-Tg between bound and free fractions. b Concentration of free, i.e. unbound thyroglobulin. This is measured in the supernatant after precipitation of IgG-bound Tg. Then, bound Tg=B:F x (free Tg) and total Tg=(bound Tg)+(free Tg). Critical factors examined in the validation of the modified method were: (1) use of immunologically intact 125I-Tg (specific activity less than or equal to 5 mCi/mg); (2) equilibration of tracer Tg with endogenous serum Tg, and (3) clean separation of IgG-bound and free fractions of Tg with goat anti-human IgG. Recovery of added Tg in the presence of TgAb was 105% and interassay precision of total Tg assay was less than or equal to 9.3%. In euthyroid subjects without TgAb, the correlation between total Tg levels by the modified and standard double-antibody method was 0.931. The correlation was much poorer in the presence of TgAb (r=0.752) reflecting their interference in the standard method. Reference ranges for serum total Tg levels by the modified method in euthyroid, hyper- and hypothyroid subjects with and without TgAb are detailed.

Clinical Utility of Thyroglobulin Antibody (TgAb) Measurements for Patients with Differentiated Thyroid Cancers (DTC)

“…This problem is exacerbated when the Tg concentration added is greater than that of the Tg in the specimen, leading to an excess of free Tg that would cause an overestimate of recovery (5, 54). Lastly, it takes time (ϳ24 h) for large molecules like Tg to form immune complexes that reach equilibrium (107,126). When a quantity of Tg is added to the specimen immediately before adding the assay reagents, there is insufficient time for the added Tg to equilibrate with the Tg and TgAb in the specimen.…”

Serum Thyroglobulin—Its Measurement and Clinical Use

“…Interference by anti thyroglobulin antibodies can be overcome either by using a second antibody (anti-rabbit IgG) which has very little cross reaction with human IgG9or by using a modified RIA. 10 These steps are time consuming and, as in the case of Black et al,9 the antibody may be difficult to obtain.…”