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Title: The development of a luminescence bioassay for thyroid stimulators
Author: Wong, Yerh Siu
ISNI:       0000 0001 3571 6832
Awarding Body: University of London
Current Institution: University College London (University of London)
Date of Award: 2001
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Graves' disease (GD) in an autoimmune thyroid disease caused by thyroid stimulating antibodies (TSAb). These mimic thyroid stimulating hormone (TSH) and result in hyperthyroidism. It has long been anticipated that an ability to quantitate TSAb in the circulation would improve the management of Graves' patients. However, to date, no entirely suitable bioassay has been developed. We aimed to exploit recent and ongoing advances in both reporter and luminescence technologies to obtain a TSAb bioassay better suited for such clinical use. Ideally, it should be precise, sensitive, rugged and technically amenable so that it has a high sample throughput. The bioassay was based upon target cells transfected to express both the human TSH receptor and also a cAMP-responsive luciferase reporter gene. An initial attempt to transfect JP26 CHO cells ourselves was unsuccessful. We therefore utilised the CHO25LUC cells which had been so transfected during the course of our work, at N.I.H. These were used for a succession of 4 protocols (Bioassays A - D) aimed at progressively refining bioassay performance characteristics. The optimal within-assay precision of Bioassay D was ~3%, as demonstrated with imprecision profiles. This was achieved by a strategy of maximizing the magnitude of bioassay response and minimizing replicate errors. Bioassay conditions needed, however, to be strictly controlled to minimize between-assay variation, since the bioassay is not rugged. We exploited the favourable bioassay precision to demonstrate unequivocally that TSH and different TSAbs do not yield parallel dose-response curves. We subsequently, adapted the bioassay for use with unfractionated human sera (10[mu]l) and finally, exploited the high throughput of the optimized bioassay in a preliminary screening of sera from Graves' patients (n = 231), comparing the results against relevant control groups.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Thyroid gland