A rat granulosa cell plasminogen activator bioassay for FSH in human serum.
A previously described in-vitro rat granulosa cell plasminogen activator bioassay for FSH has been modified and applied in the assay of human serum. This modified method consists of exposing the diethylstilboestrol-stimulated granulosa cells from 25- to 26-day-old rats to FSH or test substance for 3.5 h in wells coated with 125I-labelled fibrinogen and treated with thrombin. Following stimulation with FSH, the dose-related production of plasminogen activator was measured as the degree of 125I-labelled fibrinolysis in the presence of added plasminogen. Using the urinary FSH/LH bioassay reference preparation as the assay standard, the useful range of the assay was 0.3-15 IU/l, with an assay sensitivity of 0.3 IU/l. As determined using purified glycoprotein hormone preparations, the assay was highly specific for FSH. The minor degree of FSH bioactivity measured in some of the hormone preparations was accounted for by the amount of FSH contamination in these preparations. To abolish interference caused by unknown serum factors, we heat-treated the serum samples for 15 min at 56 degrees C before the assay. The results indicated that neither immunoreactivity nor bioactivity was affected by this treatment. Furthermore, heat-treated human sera gave responses parallel to the standard curve at the three dose levels (2, 4 and 8 microliters) studied. We used this bioassay to estimate the FSH-like bioactivity in 15 human serum samples. The estimates of immunoreactive FSH in these samples correlated well with the corresponding FSH bioactivity (r = 0.745, n = 15 and P less than 0.05). The results indicate that with this sensitive and rapid (completed within 24 h) bioassay, it should be possible to measure FSH bioactivity in heat-treated human serum samples.
Thakur
A N
AN
Department of Molecular Endocrinology, University College and Middlesex School of Medicine, London.
Coles
R
R
Sesay
A
A
Earley
B
B
Jacobs
H S
HS
Ekins
R P
RP
D000818:Animals; D001681:Biological Assay / Q000379:methods; D005260:Female; D005640:Follicle Stimulating Hormone / Q000097:blood*; D006107:Granulosa Cells / Q000378:metabolism*; D006358:Hot Temperature; D006801:Humans; D010960:Plasminogen Activators / Q000096:biosynthesis*; D051381:Rats; D011919:Rats, Inbred Strains
D005640:Follicle Stimulating Hormone; D010960:Plasminogen Activators