A water-soluble fragment of the thyroid-stimulating hormone receptor which binds both thyroid-stimulating hormone and thyroid-stimulating hormone receptor antibodies.

Previous studies have shown that freezing and thawing of human thyroid homogenates releases a water-soluble substance which reversibly binds to TSH-receptor antibodies. This substance has been designated long-acting thyroid stimulator absorbing activity (LAA). We now describe a new method for measuring LAA based on the TSH-receptor assay and application of the technique to the study of LAA. Our results indicate that LAA is a heat-labile glycoprotein which co-elutes with haemoglobin on gel filtration. Furthermore, LAA is retarded by columns of Sepharose-TSH but not by Sepharose coupled to human chorionic gonadotrophin, normal immunoglobulin G or bovine serum albumin, suggesting that LAA contains a binding site for TSH as well as for TSH-receptor antibodies. It would seem therefore that LAA is a water-soluble fragment of the TSH receptor possibly resulting from proteolytic cleavage of the receptor at a site close to the cell surface.

[Studies on the purification and the immunological properties of LATS-immunoglobulin (author's transl].

It has been shown that LATS activity is mainly distributed in the fraction of immunoglobulin G (IgG) in the serum from hyperthyroid patients. The present paper examined the immunological character of LATS and the method for separation of LATS activity from LATS positive serum using DEAE-Cellulose and affinity chromatography methods. LATS activity was distributed in the IgG fraction that could be separated by the DEAE-Cellulose column equilibrated with a 0.0175 M prosphate buffer, pH 6.3 from LATS positive serum. When LATS positive serum was fractionated by affinity chromatography on a Sepharose-bound antibody against human IgG, Fab of IgG and Fc of IgG, LATS activity was always retained in IgG fraction. When LATS positive serum was fractionated by affinity chromatography on a Sepharose-bound anti-K chain, LATS activity was found in the fraction that reacted with the anti-K chain. Because of the low antibody titer of the anti-lambda chain, LATS activity did not react with this antibody. By affinity chromatography on Sepharose-bound Concanavalin A, serum LATS activity was also retained in IgG fraction. LATS activity could be separated from LATS positive serum without significant loss of biological activity by affinity chromatography. When IgG (1) was purified from the fraction by affinity chromatography on anti-IgG (1)-bound Sepharose, about 80% of the original LATS activity was found in IgG (1) fraction. When the Fab fragment of IgG (1) was separated from papain hydrolysed IgG (1), using a Protein A-bound Sepharose column, a short-acting type of thyroid stimulating activity was found in only this fraction. These data suggest that the biological activity of the thyroid stimulation is distributed mainly in the Fab fragment of IgG (1).

Do thyroid-stimulating immunoglobulins cause non-toxic and toxic multinodular goitre?

The prevalence of serum thyroid-stimulating immunoglobulins (T.S.I.) in a variety of thyroid diseases was determined in 96 patients and 35 normal controls. Significantly elevated levels of T.S.I. were found not only in patients with Graves' disease and Hashimoto's thyroiditis but also in those with non-toxic and toxic multinodular goitre, whereas patients with a single autonomously functioning thyroid nodule, with subacute thyroiditis, and with "hyperthyroiditis" had levels which did not differ from those in the controls. We postulate that non-toxic multinodular goitre, like Graves' disease, may result from increased circulating T.S.I., which in some cases may be present in sufficient concentration to cause thyrotoxicosis.

Methods for separation of long-acting thyroid stimulator (LATS) from sera containing LATS and TSH.

Methods were devised for separation of long-acting thyroid-stimulator (LATS) from TSH in serum containing both thyroid stimulators by using Rivanol, concanavalin A (con A), or staphylococcal protein A. When 3-5 volumes of 0.5% Rivanol solution were mixed to serum containing TSH or LATS activity, LATS activity remained mainly with IgG in the supernatant fraction. On the contrary, TSH activity was precipitated. When 10 mg con A was added to 1 ml test serum, almost all TSH activity was precipitated, but LATS activity remained in the supernatant fraction, which consisted mainly of IgG and albumin. Almost all LATS activity and part of the TSH activity were precipitated by addition of more than 7.5% polyethylene glycol (PEG), which was therefore not useful for separation of the stimulators in serum. Affinity chromatography on staphylococcal protein A-Sepharose was also found to separate the two thyroid stimulators in serum. By this method LATS-immunoglobulin bound to the protein A column, but no binding of the biologic and immunologic activity of TSH was observed. The protein A method seems the most useful of these four methods for complete separation of both stimulators.

Relation of thyroid-stimulating immunoglobulins to thyroid function and effects of surgery, radioiodine, and antithyroid drugs.

Thyroid-stimulating immunoglobulins (T.S.I.) have been detected in the serum of all patients with untreated Graves' disease, and in these patients the levels of T.S.I. correlated significantly with the early uptake of 131I by the thyroid. The frequency of T.S.I. in patients treated solely by antithyroid drugs, by radioiodine, or by partial thyroidectomy was 53 per cent, 50 per cent, and 17 per cent, respectively. The reduced frequency of T.S.I. in the serum of patients treated by drugs or radioiodine was probably due to spontaneous remission, but in the case of partial thyroidectomy the operation itself clearly had a dramatic effect on the serum-T.S;I. These results accorded well with the reported frequency of thyroid autonomy in similar groups of patients and suggested that thyroid-stimulating immunoglobulins were responsible for hyperthyroidism in Graves' disease.