The radioimmunoassay of free thyroxine (T4) is a competition assay based on the principle of labeled antibody.

Samples and calibrators are incubated with 125I-labeled monoclonal antibody specific for T4, as tracer, in the presence of a biotinylated analog of thyroxine (ligand) in avidin-coated tubes. There is competition between the free thyroxine of the sample and the ligand for the binding to the labeled antibody.

The fraction of antibody complexed with the biotinylated ligand binds to avidin-coated tubes.

After incubation, the content of tubes is aspirated and bound radioactivity is measured.

A calibration curve is established and unknown values are determined by interpolation from the curve.

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