Comparisons of Different Assays for the Thyroid-Stimulating Antibody of Graves’ Disease*

Abstract

Various current methods for the assay of the thyroid-stimulating antibody of Graves' disease (TSAb) were assessed for comparative sensitivity and specificity. Five procedures were examined in detail: I) increase in cAMP in human thyroid slices; II) inhibition of [125I]TSH binding to human thyroid membranes, particulate or III) solubilized; IV) inhibition of [125I]TSH binding to guinea pig fat cell membranes, particulate or V) solubilized. The minimum effective concentration of TSH (microunits per ml) in these assays was: I) 5; IV) 12.5; V) 25 and II and III) 100. The guinea pig fat systems (IV and V) were more sensitive than the human thyroid assays (II and III) in terms of the concentration of TSH required for 50% inhibition of binding. Five preparations of TSAb-immunoglobulin G were found in general to have constant relative potency in the five assay systems, but individual immunoglobulins G were variable in their effects when data in the five procedures were compared directly. It is probable that, in addition to TSAb, other antibodies, more or less specific for the human thyroid, influenced the TSH binding inhibition assays to a variable extent. The above TSH binding inhibition assays were carried out with solutions or suspensions of receptor preparations. Solid phase systems, using microtiter plates for TBI assays, were assessed in less detail. Human thyroid, particulate or solubilized, was ineffective (little specific binding of [125I]TSH), but a particulate preparation of guinea pig fat cell membranes was an efficient basis for a solid phase TBI system. Solid phase assessment with [125I]staphylococcal protein A of binding of thyroid autoantibodies to human thyroid membranes was not specific for TSAb.

A predicted case with neonatal transient hypothyroidism due to blocking type thyrotropin binding inhibitor immunoglobulins (TBII)

“…Recently, TSH receptor related antibodies have become measurable by precise in vitro radioassays (Zakarija et al, 1980;Sugenoya et al, 1979;Loeffler et al, 1983). Most immunoglobulins from untreated Graves' patients competitively inhibit the binding of 125I-TSH to the TSH receptor, and these immunoglobulin activities have been reported to correlate well with thyroid function.…”

“…If potent TBII of the blocking type is present in the maternal blood, then it should cause neonatal hypothyroidism by its placental passage as is the case of neonatal Graves' disease (Zakarija et al, 1983).…”

Properties of thyrotropin (TSH) binding to plasma membranes prepared from rat fat tissues.

“…In the present study we used the same labeled bTSH and incubation buffer to determine the TSH receptors on rat fat membranes, so a single class of binding sites was thought to be specific to rat fat membranes. The value of Ka of rat fat membranes was lower than that of high affinity site of human thyroid membranes, although the values of guinea pig fat membranes were reported to be similar to that of human thyroid membranes (Endo et al 1981;Loeffier et al 1983).…”

“…Kishihara et al (1981) reported that a single class of binding site for TSH could be observed in guinea pig fat membranes using highly purified labeled bTSH although with relatively crude labeled bTSH two binding sites were detected. On the other hand, Loeffier et al (1983) reported two binding sites for TSH using highly purified TSH. Other reports suggested that ionic strength of incubation buffer affected TSH binding to thyroid membranes (Powell-Jones et al 1980; Pekonen and Weintraub 1980).…”

Acute exogenous TSH administration stimulates leptin secretion in vivo

“…High affinity TSH binding activity of Graves' IgG has been shown in guinea pig adipocytes (2)(3)(4), and Tshr cDNA fragments have been obtained by PCR from rat or human retro-orbital tissues, adipose tissue, and fibroblasts (5)(6)(7)(8). In vitro studies indicate that TSH-mediated responses on adipocytes include proliferation, differentiation, survival, and stimulation of specific cellular pathways (9)(10)(11)(12)(13)(14)(15)(16)(17).…”