Circulating Thyroglobulin-Antithyroglobulin Immune Complex in Thyroid Diseases Using Enzyme-Linked Immunoassays

Abstract

Highly sensitive, solid phase enzyme-linked immunoassays were developed for the determination of circulating thyroglobulin-antithyroglobulin immune complexes (Tg • anti-Tg IC). In an assay using β-D-galactosidase conjugated with the monovalent fragment (Fab') from rabbit (anti-Tg) immunoglobulin G (IgG) and silicone rods coated with rabbit (antihuman IgG) IgG, the minimal detectable amount of Tg•anti-Tg IC was as small as 0.04 ng⁄tube, corresponding to approximately 10 ng⁄ml serum. In another assay using the same enzyme conjugate and silicone rods coated with protein A from Staphylococcus aureus, the smallest amount of detectable Tg•anti-Tg IC was 0.04 ng⁄ tube, corresponding to 3 ng⁄ml serum. The specificity of these assays was demonstrated by 1) parallelism of the standard curve with dilution of serum samples from patients with thyroid diseases, and 2) nondetectability of Tg•anti-Tg IC in the sera of normal subjects. Coefficients of variance within and between assays for the two measurement systems were 9.8–15.6‰ and 9.9–21.2‰, respectively, in the first assay and 8.2–14.5‰ and 16.5–25.7‰, respectively, in the second assay. Tg•anti-Tg IC was detectable in 4 of 14 patients with Graves' disease (29‰) and in 3 of 20 patients with Hashimoto's thyroiditis (15‰) using the first assay system. Levels were detectable in 7 of 15 patients with Graves' disease (47‰) and in 12 of 37 patients with Hashimoto's thyroiditis (32‰) using the second assay system. The circulating levels of Tg•anti-Tg IC ranged from 8.7–498 ng⁄ml. Tg•anti-Tg IC was more frequently detectable in sera containing more than 10 ng⁄ml Tg and less than 10 μg⁄ml anti-Tg; these conditions were more frequently observed in sera from patients with Graves' disease (Tg, 67 ± 144 ng⁄ml; anti-Tg, 1.4 ± 2.5 μg⁄ml; mean ± SD; n = 55) than in sera from patients with Hashimoto's thyroiditis (Tg, 26 ± 73 ng⁄ml; anti-Tg, 16.4 ± 31.4 μg⁄ml; mean ± SD, n = 120). Three of 11 patients with Graves' disease and detectable IC in their sera had associated ophthalmopathy.