Evaluation of T4 radioimmunoassay as a screening test for thyroid function

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Various tests have been developed for thyroid function based on the measurement of total serum thyroxine (T4) level, either by direct or indirect techniques. The direct assays include competitive protein binding assay and radioimmunoassay. Radioimmunoassay of T4 provides distinct advantages over competitive protein binding assays by eliminating extraction procedures and using a smaller amount of serum. Of the indirect assays, the most commonly used screening test for thyroid function is the commercially available in vitro blood test, effective thyroxine ratio (ETR).

The present studies describe the development and clinical evaluation of a double antibody radioimmunoassay for serum thyroxine and its comparison with ETR in different thyroid disorders.

Materials and methods

Production of antiserum. Antiserum was produced in rabbits by immunizing with T4 conjugated to bovine serum albumin (BSA) using the method for triiodothyronine (T3) antibodies described by Gharib et al.1 Four rabbits were immunized weekly with 1 mg antigen; detectable antibodies were produced in 4 weeks and adequate titers in 12 weeks. Antiserum from rabbit 1 (titer 1:5000) was used for these studies.

Preparation of T4 free serum. T4 free serum was prepared by removing exogenous T4 with charcoal adsorption. Normal human serum (100 ml), to which 10 g charcoal (Norit A) was added, was stirred overnight (24 hours) at 4C. Then the charcoal was removed by centrifugation: it was centrifuged three times at 13,000 rpm for 1 hour each. The process removed over 99% of added T4 125I without making significant change in total protein concentration.

Eight-anilino naphthelene sulfonic . . .



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