Clinical studies on autoimmune mechanisms of the thyroid. Part 2. Studies on measurements of serum thyroblobulin concentrations in patients with various thyroid diseases (author's transl)

Abstract

The present report deals with the measurement of serum thyroglobulin (Tg) in various thyroid disorders by using radioimmunoassay (RIA). The results were as follows: 1) A specific and simple solid-state RIA for the measurement of Tg in human serum was used. This system was a direct RIA using plactic cups coated with crude anti-thyroglobulin antibodies (anti-Tg) and 125I labeled purified anti-Tg. The purification of anti-Tg was performed by affinity chromatography using Tg-Sepharose, as an immunoadsorbent. 2) Affinity chromatography was carried out using a modification of the method of Cuatrecasas. The immunoadsorbent (Tg-Sepharose conjugate) was used in a column procedure for the isolation of anti-Tg from globulin fractions obtained from Hashimoto's sera. The elution was performed with 4M NaI or 0.17M Glycine-HCl with pH 2.3. The eluted materials contained a very small amount of Tg which was removed by Sephadex G-200 chromatography using the same elution buffer. The high purity of the anti-Tg obtained was demonstrated by the fact that almost all of the final product was bound with Tg by using Sepahdex G-200 chromatography. 3) A direct RIA consisting of two incubation steps was applied for Tg measurement. During the first incubation, standard Tg or Tg in serum was bound to the antibody coated cup. After washing, equal amounts of radioactive purified anti-Tg were incubated with the bound Tg. The cups were then washed again, and counted separately. The radioactive counts thus obtained, increased with the amounts of Tg bound to the anti-Tg by the first incubation. The sensitivity of the assay was 4 ng/ml. T3 and T4 did not cross-react against Tg, and did not interfere with the binding between Tg and 125I-anti-Tg. A dilution curve was constructed using the serum of a patient with Graves' disease; the post-operative serum contained a very high level of Tg. This curve paralleled the standard curve. By adding constant Tg to a normal serum or Graves' serum, Tg recovery was good. However, in regard to Hashimoto's serum, Tg recovery was unsatisfactory. 4) Serum Tg concentrations were measured in patients with various thyroid disorders. ..