We have assessed a new method of free T4 measurement (Amerlex) which uses a novel unidentified T4-labelled analogue, said to be unreactive with T4 binding proteins in serum, together with an antibody that binds both analogue and T4. Free T4 is assessed by competition with analogue for antibody binding-sites. The test method has been compared with free T4 measured by equilibrium dialysis and with a technique using an immobilized T4 antibody. All methods gave the expected free T4 levels in normal, hyperthyroid and hypothyroid subjects and normal free T4 levels with high or low levels of T4 binding globulin. However, in autosomal dominant familial euthyroid T4-excess, where T4 is abnormally bound to albumin, the test method gave apparent high free T4 levels suggestive of hyperthyroidism. In a selected group of severely-ill euthyroid patients the new method gave apparent low free T4 levels. In view of these discrepancies, binding of labelled analogue was evaluated by dextran-charcoal separation of 4 degraees C. Familial euthyroid T4-excess sera showed greater analogue binding and samples with low prealbumin concentration showed less binding than did normal sera. Despite its validity with variations in TBG, it appears that Amerlex Free T4 is influenced by lower-affinity, high-capacity T4 binding sites in serum, so that apparent free T4 concentration may vary with changes in the concentration of such sites.

The abnormal high capacity T4 binding site of familial euthyroid T4 excess was separable from prealbumin and T4-binding globulin but not from albumin. We therefore compared T4 binding by albumin preparations isolated from the sera of normal and affected subjects. By equilibrium dialysis, albumin from affected subjects showed an extra T4 binding site (Kd approximately 50 nM) in addition to the T4 binding sites of normal albumin (Kd approximately 4 microM). Comparison of the estimated capacity of the additional site (200 microM) with the molar concentration of albumin suggested that only about one third of albumin molecules from affected subjects contained the extra binding site. Estimates of affinity and capacity were used to derive combining powers for the diverse classes of serum T4 binding sites. From these estimates, it appears that the presence of the abnormal site accounts for the approximate doubling of normal mean total T4 (from approximately 100 nM or 7.7 micrograms/dl to approximately 200 nM or 15.5 micrograms/dl), in order to maintain a normal free T4 in the face of the increased T4 association with albumin. Studies of [125I]T4 displacement from albumin of affected subjects showed low T3 affinity and competition by barbitone. Relative molar concentrations to give equivalent displacement of [125I]T4 were: 3,3',5,5'-tetraiodothyroacetic acid, 0.4; T4, 1.0; rT3, 4; 8-anilinonaphthalene sulfonic acid, 10; T3, 80; salicylate, 200; and barbitone, 40,000. Studies with dithiothreitol suggested that disulfide bonds were critical in maintaining the T4-albumin association. These findings indicate that familial T4 excess is due to abnormal intermediate affinity, sulfhydryl-sensitive T4 binding sites that are inseparable from the albumin found in affected subjects.

1. Immunoreactive angiotensin II was measured in cerebrospinal fluid of four normal dogs. 2. The migration of this immunoreactive angiotensin II on polyacrylamide-slab gel electrophoresis was identical with the migration of the heptapeptide, Des-Asp1-angiotensin II, in each case. 3. The biological activity of the material from canine cerebrospinal fluid in a pressor bioassay was similar to that of Des-Asp1-angiotensin II. 4. The pressor activity of the canine material was abolished by treating the pressor bioassay rat with a competitive antagonistic analogue, Sar1-Ala8-angiotensin II. 5. The results suggest that the biologically active immunoreactive angiotension II present in normal canine cerebrospinal fluid is composed mainly of the heptapeptide fragment of angiotensin II, Des-Asp1-angiotensin II.

Angiotensin converting enzyme was measured in the serum of 52 patients with sarcoidosis, 67 healthy control subjects and 64 patients with pulmonary and non-pulmonary diseases. The patients with sarcoidosis were classified on clinical grounds as having active or inactive disease. In 26 patients with active sarcoidosis not taking corticosteroids the mean serum ACE was significantly higher than in normal controls (P less than 0 . 001). 73% of these patients had elevated serum ACE. Only two out of 12 (17%) patients with inactive sarcoidosis not taking corticosteroids had elevated serum ACE. Serum ACE was normal in patients taking oral corticosteroids for longer than two weeks. Eighty per cent of patients with active sarcoidosis with radiological evidence of pulmonary parenchymal involvement had an elevated serum ACE compared to 25% in patients with normal chest X-rays and 60% of those with bilateral hilar lymphadenopathy. All sarcoid patients with hypercalcaemia had elevated serum ACE whereas only half of those with normal serum calcium had elevated ACE. In the patients with other thoracic and granulomatous conditions serum ACE was normal or rarely marginally elevated. Serum ACE appears to be of value in the diagnosis of active sarcoidosis.