A Method for the Quantitative Determination of Gonadotrophins in the Urine of Non-Pregnant Human Subjects

Abstract

A method for the quantitative determination of gonadotrophins in the urine of non-pregnant human subjects is described.The reliability criteria for the method have been studied and the technique has been found to be satisfactory in terms of accuracy, precision, sensitivity and specificity.The importance of expressing results of urinary gonadotrophin assays in terms of a reference preparation such as HMG 20A is emphasized.The aim of the present paper is to describe a quantitative method for the estimation of gonadotrophins in urine of non-pregnant human subjects and to evaluate the method in terms of accuracy, precision, specificity and sensitivity. The method essentially consists of adsorption of the gonadotrophins on to kaolin, elution from kaolin, precipitation by acetone and, in the case of low-titre urines, treatment of the crude kaolin-acetone powders by tricalcium phosphate. The bioassay is performed using the mouse uterus test and results are expressed in terms of a standard prepared from the urine of menopausal and postmenopausal women and termed HMG 20A. The preliminary work on which the method is based has already been described [Loraine & Brown, 1954Loraine, 1957Loraine, , 1958a. MATERIALS AND METHODSThe kaolin was acid washed (Society of Leather Trades Chemists' specification) and was supplied by British Drug Houses, Ltd. The tricalcium phosphate suspension was prepared according to the method of Swingle & Tiselius [1951] and contained approx. 0-07 g Ca3(P04)2/ml. Hydrogen-ion concentrations were measured by means of a Cambridge pH meter using a glass electrode and were uncorrected for ionic concentration.The standard (HMG 20 A) was prepared by Messrs Organon (Newhouse, Scotland) from the urine of menopausal and postmenopausal women. The kaolin-acetone method was used. The crude powder thus obtained was treated with tricalcium phosphate, subjected to fractional acetone precipitation and then dried over phosphorus pentoxide. A 'unit' was defined as the gonadotrophic activity contained in 1 mg HMG 20 A [Report of the second meeting of the Gonadotrophin Club, 1955, unpublished].

Expression of human choriogonadotropin in monkey cells using a single simian virus 40 vector.

“…The biological activity of the rhCG dimer produced in CV-1 cells was determined in vivo by the mouse uterine weight assay (19). The increase in weight and size of the uteri with respect to those of the untreated female mice is a measure of biologically active hCG.…”

“…The biological activity of the hCG produced by the CV-1 cells was determined in vivo by the mouse uterine weight assay (19). Two milliliters of tissue culture supernatants was injected (i.p.)…”

Amenorrhoea in the Feeding Disorders: Anorexia Nervosa and Obesity

“…Total gonadotrophin activity (TGA) in the urine was estimated by the method of Loraine and Brown [1959]. This is a bioassay which is non-specific inasmuch as it is total gonadotrophin activity which is measured, and it is not possible to distinguish be tween the relative proportions of FSH and LH.…”

Psychological and nutritional factors in disturbances of menstrual function and ovulation

“…Twenty-four-hour collections of urine were made serially over periods ranging from 50 to 120 days. In the main studies, total gonadotrophic activity (TGA) in the urine was measured by the bio-assay method of Loraine & Brown (1959). This is a bio-assay which is non-specific in as much as it measures total gonadotrophic activity and cannot distinguish between the relative proportions of follicle-stimulating hormone (FSH) and luteinizing hormone (LH).…”