Development of a sensitive porcine granulosa cell bioassay for follicle-stimulating hormone (FSH).
OBJECTIVE
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.


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.
Sites
C K
CK
Department of Obstetrics and Gynecology, University of Vermont College of Medicine, Burlington 05405, USA.
Taylor
E H
EH
Lewis
S J
SJ
Brumsted
J R
JR
D000735:Androstenedione / Q000031:analogs & derivatives / Q000378:metabolism; D000818:Animals; D001141:Aromatase / Q000378:metabolism; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004958:Estradiol / Q000378:metabolism; D005260:Female; D005470:Fluorometry; D005640:Follicle Stimulating Hormone / Q000032:analysis* / Q000097:blood / Q000494:pharmacology; D006107:Granulosa Cells / Q000378:metabolism*; D066298:In Vitro Techniques; D010907:Pituitary Hormones / Q000494:pharmacology; D051381:Rats; D013552:Swine
D010907:Pituitary Hormones; D000735:Androstenedione; D004958:Estradiol; D005640:Follicle Stimulating Hormone; C011482:19-hydroxy-4-androstene-3,17-dione; D001141:Aromatase