|Back Topic Categories Search Previous Abstract Next Abstract|
Anita Narula1 , Petri Ryhanen2 , Pulak Manna2 , Ilpo Huhtaniemi2 , William Bremner1 *
Department of Medicine, University of Washington, Seattle, Washington 1
Department of Physiology, University of Turku, Turku, Finland 2
Effects of FSH are mediated by its interaction with a specific receptor through G-coupled proteins, resulting in increased levels of cAMP. We generated a granulosa cell line from an ovarian tumor of a transgenic mouse expressing inhibin a-promoter/SV 40-T Ag fusion gene. This cell line was stably transfected with the recombinant human FSH receptor and was used to develop an in vitro bioassay for FSH. The FSH response was estimated based on the measurement of extracellular cAMP. The cell line expresses 25 × 103 receptors/ cell, which binds FSH with high affinity (Kd: 0.69x 10-9M). A dose dependent increase in cAMP (ED50:10 mIU/mL) was observed after stimulation of cells with rec. hFSH. The sensitivity of the assay is <0.8mIU/mL. The intra- and interassay coefficients of variation are: 8.2 and 13.1%, respectively. The serum interfering factors were removed without much loss of FSH by pretreatment of samples with polyethylene glycol (12%). Postmenopausal and castrated monkey serum exhibited parallel dose-response curves. Therefore, this assay can be effectively used to measure bioFSH levels using 20ul of serum sample. This assay is highly specific for FSH and no significant increase in cAMP was discernedwith human LH, CG, TSH, PRL and GH. Monkey, rat, ovine and porcine FSH showed a parallel dose-dependent increase in cAMP production. This assay system provides an important tool in detecting the bio FSH levels in physiological and pathological conditions in humans and other species.
This abstract is being presented on Sunday, August 1 at 8:00 AM to 10:15 AM at CUB 2nd Floor Ballroom.