Development of a highly sensitive and selective microplate chemiluminescence enzyme immunoassay for the determination of free thyroxine in human serum

Xu Wang¹, Hui Chen¹, Jin-Ming Lin¹, Xitang Ying²

1. The Key Laboratory of Bioorganic Phosphorus Chemistry & Chemical Biology, Department of Chemistry, Tsinghua University, Beijing, 100084, China
2. Beijing Chemclin Biotech Co., Ltd., Beijing Academy of Science and Technology, Haidian, Beijing, 100094, China

Abstract

A microplate chemiluminescence enzyme immunoassay (CLEIA) with high sensitivity, selectivity and reproducibility was developed for the determination of free thyroxine (FT4) in human serum. A competitive assay has been utilized with horseradish peroxidase (HRP) labeled thyroxine analog in the chemiluminescence (CL) detection. The CL signal produced by the emission of photons from luminol was directly proportional to the amount of analyte. The linear range was 0.45-7.5 ng dL^-1and the detection limit was 0.09 ng dL^-1. Experimental conditions, such as temperature, pH, incubation time, titration level and other relevant variables upon the CL signal have been examined and optimized. A coefficient of variance of less than 16% was obtained for intra- and inter-assay precision. The present method has been successfully applied to the analysis of FT4 in human serum. The positive and negative coincidence ratios are satisfactory. Good correlations were obtained between the results by the proposed method and radioimmunoassay (RIA), as well as a Bayer ACS-180SE detection system.

Keywords: Free thyroxine (FT4), horseradish peroxidase (HRP), chemiluminescence enzyme immunoassay (CLEIA)

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How to cite this article:
Wang X, Chen H, Lin JM, Ying X. Development of a highly sensitive and selective microplate chemiluminescence enzyme immunoassay for the determination of free thyroxine in human serum. Int J Biol Sci 2007; 3(5):274-280. doi:10.7150/ijbs.3.274. Available from http://www.ijbs.com/v03p0274.htm