Analysis of free hormone fractions by an ultrafast immunoextraction/ displacement immunoassay: Studies using free thyroxine as a model system

William Clarke, John E. Schiel, Annette Moser, David S. Hage

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Abstract

A system was developed for measuring the noncomplexed or free fraction of a hormone in serum based on the combined use of ultrafast immunoextraction with a chromatographic displacement immunoassay. This approach was tested using L-thyroxine as a model analyte. Items considered in the development of this technique included the choice of immunoassay format and the selection of conditions for removal of thyroxine's free fraction from samples without significant interference from its protein-bound fraction. The final method had an effective extraction time of 90 ms and allowed the amount of free thyroxine to be determined within 30 s after sample injection. The limit of detection was 6 pM (S/N = 3) for a 100-μL sample, and the linear response extended up to at least 100 pM. This technique gave good correlation versus reference methods when used for the determination of free thyroxine in serum samples. Advantages of this method included its speed and its ability to analyze a sample with no pretreatment other than standard filtration. The same approach could be adapted for other hormones or drugs by using appropriate antibodies and labeled analogues for such agents.

Original languageEnglish (US)
Pages (from-to)1859-1866
Number of pages8
JournalAnalytical chemistry
Volume77
Issue number6
DOIs
Publication statusPublished - Mar 15 2005

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ASJC Scopus subject areas

  • Analytical Chemistry

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