PURPOSE: Our objective was to develop a sensitive in vitro bioassay for follicle-stimulating hormone (FSH) that does not require the housing of animals in a research facility. MATERIALS AND METHODS: Porcine granulosa cells from 1- to 3-mm follicles were cultured on laminin for 48 hr in serum-free medium in the absence or presence of FSH or with other purified pituitary hormones, supplemented with 19-OH androstenedione. Estradiol accumulation in medium per microgram of DNA of cells was determined as a reflection of FSH-induced aromatase activity. RESULTS: FSH (0.01-10 ng/ml) caused a dose-dependent increase in estradiol production per microgram of DNA, with 1, 10, and 100 ng/ml significantly higher than control. Porcine FSH was approximately two fold more biopotent than rat FSH in this system. Higher doses of FSH (100 ng/ml) caused less estradiol accumulation, presumably reflecting FSH receptor down regulation. No other pituitary hormone produced significant estradiol accumulation. Unextracted serum from a patient with premature ovarian failure (10-50 microliters) was tested in parallel to purified rat FSH (0-50 ng/ml) in this system, resulting in similar estradiol accumulation per microgram of DNA. CONCLUSIONS: We have developed a porcine granulosa cell bioassay for FSH which is sensitive, is specific for FSH, and does not require the housing of animals on site. It can be completed by a technician within 4 working days and can detect FSH in a sample of human serum.
PURPOSE: Our objective was to develop a sensitive in vitro bioassay for follicle-stimulating hormone (FSH) that does not require the housing of animals in a research facility. MATERIALS AND METHODS: Porcine granulosa cells from 1- to 3-mm follicles were cultured on laminin for 48 hr in serum-free medium in the absence or presence of FSH or with other purified pituitary hormones, supplemented with 19-OH androstenedione. Estradiol accumulation in medium per microgram of DNA of cells was determined as a reflection of FSH-induced aromatase activity. RESULTS: FSH (0.01-10 ng/ml) caused a dose-dependent increase in estradiol production per microgram of DNA, with 1, 10, and 100 ng/ml significantly higher than control. Porcine FSH was approximately two fold more biopotent than rat FSH in this system. Higher doses of FSH (100 ng/ml) caused less estradiol accumulation, presumably reflecting FSH receptor down regulation. No other pituitary hormone produced significant estradiol accumulation. Unextracted serum from a patient with premature ovarian failure (10-50 microliters) was tested in parallel to purified rat FSH (0-50 ng/ml) in this system, resulting in similar estradiol accumulation per microgram of DNA. CONCLUSIONS: We have developed a porcine granulosa cell bioassay for FSH which is sensitive, is specific for FSH, and does not require the housing of animals on site. It can be completed by a technician within 4 working days and can detect FSH in a sample of human serum.