An enzyme immunoassay for human follicle-stimulating hormone

Abstract

An enzyme immunoassay for human follicle-stimulating hormone (hFSH) has been developed utilizing highly purified hFSH coupled to glucose oxidase (EC 1.1.3.4) as the tracer, and antiserum to hFSH. Coupling of hormone and enzyme was done in the presence of glutaraldehyde and separation of bound hormone-enzyme complex from unbound was by double-antibody precipitation. Glucose oxidase activity was measured in buffer-reconstituted pellets derived from precipitation by second antibody. The hFSH-glucose oxidase conjugate reained immunologic activity after storage for 7 months at −20°C. Follicle-stimulating hormone activity, in a series of human pituitary fractions varying in potency from 125 to 3608 IU/mg as determined by a rat ovarian weight gain bioassay, gave similar potency estimates by enzyme immunoassay, with an index of discrimination of 1.10. The enzyme immunoassay proved useful for measurement of hormone mass in preparations of radiolabeled hFSH, allowing accurate calculation of specific radioactivity. The mean index of precision λ for a series of 20 enzyme immunoassay measurements was 0.079, with a detection limit of 4 mIU, about 1 ng of purified hFSH. The enzyme immunoassay gave results in good agreement with those from radioimmunoassay when applied to measurement of follicle-stimulating hormone levels in dialyzed human serum with an index of discrimination (enzyme immunoassay/radioimmunoassay) of 1.05. Recovery of varying amounts of hFSH added to a single serum pool was greater than 90%. Since the enzyme immunoassay does not require isotopes or specialized counting equipment, it may represent an attractive alternative to radioimmunoassay for laboratories with limited facilities.