Ian H. Thorneycroft

Radioimmunoassay of serum progesterone in women receiving oral contraceptive steroids

Abstract A dextran-coated charcoal radioimmunoassay (RIA) of serum progesterone which requires only a simple diethyl ether extraction of serum prior to assay is presented. The assay uses a very specific antiserum produced by immunizing rabbits with an 11α-succinylprogesterone-bovine-serum albumin conjugate. The advantage of the method presented over other RIA methods for progesterone is that no chromatography is required. The mean serum progesterone concentrations measured were for men, 193 pg/ml; for women in the follicular and luteal phases of the menstrual cycle, 191 pg/ml and 8.9 ng/ml, respectively; for post-menopausal women, 148 pg/ml; and for ovariectomized women, 109 pg/ml. Sera were also assayed from women receiving combination oral contraceptives and their mean progesterone concentration was 309 pg/ml. It is concluded that ovarian function is not depressed to post-menopausal levels in women receiving oral steroidal contraceptive agents.

Serum gonadotropin and steroid patterns during the normal menstrual cycle

n At the University of Southern California School of Medicine, the reproductive hormones follicle-stimulating hormone (FSH), luteinizing hormone (LH), progesterone, and estradiol were measured in serum samples obtained daily from a group of women throughout a normal menstrual cycle. Competitive binding techniques were used for the analysis of aliquots from the 10 women, aged 20-28, whose hormonal levels were studied. The results were generally in agreement with those of previous investigators, whose separate researches were less extensive than the research described here. FSH showed an early follicular phase rise, a late follicular phase decline, and a midcycle peak occurring on the day of the LH peak or on the day before and followed by a luteal phase decline. LH showed a slight progressive rise in the follicular phase, a midcycle peak, and a slight fall in the luteal phase. Estradiol also reached a midcycle peak. After the midcycle peaks, a rise progesterone. Progesterone and estradiol fell a few days before menstruation.n

Single luteal phase serum progesterone assay as an indicator of ovulation

n A study was undertaken to determine whether solitary progesterone as says performed on serum samples obtained in the midluteal phase would provide the clinician with a convenient indicator that ovulation had occurred in that cycle. After a normal luteal-phase range was establish ed, single luteal-phase serum progesterone sampling was performed in 51 infertile women with regular menses and 35 oligomenorrheic women undergoing clomiphene citrate therapy. In the follicular phase of the cycle, progesterone levels were consistently less than 2 ng/ml. Between 11 and 4 days prior to the onset of menses in presumptively ovulatory cycles, serum progesterone levels were always 3 ng/ml or greater. Progesterone values in this range were always accompanied by a secretory endometrium and can be considered presumptive evidence of ovulation. This rapid, easily performed technique enables 1 technician to assay 30 or more samples for progesterone in a single working day and the results are available within 24 hours. This assay technique is easier to perform and more reproducible than a urinary pregnanediol assay, and it is expected that clinical laboratories will soon perform serum progesterone assays as a routine procedure.n

Serum testosterone concentrations in women throughout the menstrual cycle and following HCG administration.

Abstract Serum testosterone (T) concentrations, measured by a specific, precise, and sensitive radioimmunoassay in 40 women with apparently normal menstrual cycles averaged 34.6 ± 10.3 (S.D.) ng. per cent and ranged from 14 to 59 ng. per cent. Mean serum T concentrations, assayed daily in eight women throughout an entire ovulatory cycle, were highest around the midcycle LH peak and higher during the follicular than during the luteal phase of the cycles. Averages of daily serum T concentrations, when determined for each individual woman for the entire cycle, varied significantly among the eight subjects studied, ranging from 23.1 to 39.0 ng. per cent. Coefficients of variation of serum T levels in these eight individual cycles averaged 22 per cent. HCG administration to three of these eight women during the luteal phase of their subsequent cycle resulted in a significant rise of serum T concentrations in only one subject. These data indicate that mean serum T concentrations in normal women are subject to small but significant changes during the course of the menstrual cycle. Despite these cyclic changes, as well as day-to-day variations, serum T concentrations in normal women largely fall into a relatively narrow range. Repeated serum T concentrations which consistently exceed 55 ng. per cent (2 S.D. above the mean) may be regarded as documentation of androgen excess.

The relation of serum 17-hydroxyprogesterone and estradiol-17β levels during the human menstrual cycle ☆

Abstract Concentrations of 17-hydroxyprogesterone (17-OHP) were measured in serum samples obtained daily through 9 menstrual cycles. These samples had previously been assayed for estradiol, progesterone, luteinizing hormone (LH), and follicle-stimulating hormone. The concentration of 17-OHP increased at mid-cycle and continued to be high throughout the luteal phase of the cycle. The first sustained rise in 17-OHP levels was associated with the initiation of the mid-cycle LH surge. The results of this study indicate that the mid-cycle rise of 17-OHP may well be one of the earliest indicators of luteinization of the follicle as this hormone appears to be initially secreted by luteintzed thecal cells and then by the corpus luteum. It is concluded that estradiol levels rather than 17-OHP levels provide a good index of follicular maturation, whereas rising concentrations of the latter hormone indicate luteinization of the follicle.

A radioimmunoassay of androstenedione.

Abstract A dextran-coated charcoal radioimmunoassay for androstenedione (4-androsten-3, 17-dione) is reported which uses an anti-testosterone antiserum raised in sheep, against a testosterone-17-hemisuccinate-Bovine Serum Albumin conjugate. It is more sensitive and rapid than previously published double dilution, gas chromatographic and competitive protein binding assays. Androstenedione is separated from cross-reacting Steroids by Celite column chromatography. The intra-assay and interassay coefficients of variation were 10.7 and 11.6 per cent, respectively. Using this method serum androstenedione in men was 1.15 ± 0.35 ng/ml; in women, 1.41 ± 0.30 ng/ml; in post-menopausal women, 0.88 ± 0.34 ng/ml; in ovariectomized women, 0.67 ± 0.17 ng/ml; and in ovariectomized-adrenalectomized women, 0.14 ± 0.05 ng/ml. The blank of the method was usually 4 to 5 pg, but ranged between 0 and 12 pg. The sensitivity of the standard curve was 8 pg.

Serum gonadotropin and steroid patterns in early human gestation

n Estradiol (E), progesterone (P), 17-hydroxyprogesterone (17-P), follicle stimulating hormone (FSH), luteinizing hormone-human chorionic gonadotropin (LH-HCG), and human placental lactogen (HPL) concentrations were measured in serum samples obtained daily from 3 women from the last menstrual period (LMP) throughout the 1st few months of gestation. Radioimmunassay of serum samples was used. FSH levels declined after implantation in 1 subject and remained unchanged in the others. HCG levels began to rise above luteal phase LH values 11-14 days after the midcycle LH peak. HPL became detectable 34-38 days after the LH peak. E levels rose steadily after the postovulatory nadir, increasing rapidly 4 weeks after the LH peak. Patterns of P and 17-P were similar initially; both rose after ovulation, remained elevated for several weeks, and then declined. About 11 weeks after the LMP, levels of P again increased whereas 17-P remained low. Single serum samples were obtained from an additional 158 gravid women 6-16 weeks after the LMP, and E, P, and 17-P were measured in each. Mean levels of E steadily increased and 17-P steadily decreased. Mean P dropped to a nadir at 9 weeks and rose therafter. Since the 17-P is mainly of luteal origin, these findings indicated that in normal pregnancy the corpus luteum has maximal activity for about 4 weeks after ovulation. Falling levels of P in the 3 weeks prior to increased trophoblastic production of this hormone may contribute to the cause of 1st trimester uterine bleeding in some women who have normal term pregnancies.n

Log normal distribution of gonadotropins and ovarian steroid values in the normal menstrual cycle

In the statistical analysis of the values of luteinizing hormone, follicle-stimulating hormone, estradiol, and progesterone obtained from normal menstrual cycles, a depature from normality was noted. Chi square, W test, and linear transformation were used to check the normality of the distributions. The results of this investigation showed that the distributions were not normal (Gaussian) but log-normal. By plotting the probit of the percentages of cumulative frequency on a log scale (probit-log), linearity of the data was obtained. This resulted in direct graphical estimations of values with a useful clinical range, which included the mean and the 95 per cent confidence interval.

Estrogenic activity in women receiving an injectable progestogen for contraception

n 121 multiparous women (aged 15-37) who had received injections of 150 mg of depomedroxyprogesterone acetate (DMPA) every 3 months as a contraceptive for more than 1 year, were examined to assess possible clinical alterations due to low estrogen levels. Serum samples were taken to evaluate estradiol levels by radioimmunoassay. Clinical examination did not reveal any sign of estrogen deficiency, except a smaller uterine than expected. Serum estradiol concentrations (36-42 pg/ml) were in the range of the follicular phase estradiol levels of the normal ovulating woman. As this follicular level probably persists throughout treatment, total estrogenic stimulation of end organs was much lower than for cycling women, but greater than for post-menapausal women. Patients receiving DPMA have functioning ovaries which continue to secrete estradiol at follicular levels. Uterine atrophy and shift in maturation index during contraceptive treatment with this drug is due to the large dose of progestogen outweighing the low levels of estradiol. It is suggested that long-term studies are needed to determine the possible advantage of increasing the estrogen effect on end organs over the known adverse effects of estrogen administration.n

Clinical categorization of patients with secondary amenorrhea using progesterone-induced uterine bleeding and measurement of serum gonadotropin levels

A group of ninety unselected women with secondary amenorrhea of at least six months duration were studied retrospectively. By the use of intramuscular progesterone in oil, it was possible to categorize these patients according to their positive or negative uterine bleeding response. LH, FSH, and estradiol values in the 63 patients of the positive category had a log-normal distribution. LH values were found to be composed of two different populations; FSH and estradiol values were composed of only one population. Based on these LH determinations the entire positive category was then divided into two groups. Patients with high levels of LH (Group I) were clinically diagnosed to have polycystic ovarian disease. Patients with normal levels of LH (Group II) were diagnosed to have hypothalamic-pituitary dysfunction. The 27 patients who failed to have withdrawal uterine bleeding were placed in the negative category. These patients also were shown to have a log-normal distribution for LH, FSH, and estradiol. In contrast to the patients in the positive category, FSH values in these patients were made up of two different populations whereas LH and estradiol values showed only one population. Based on the FSH values these patients were divided into two groups, one with low and normal levels of FSH (Group III) and the second population with high levels of FSH designated as Group IV. Group III represented those patients with hypothalamic-pituitary failure and patients in Group IV were those with ovarian failure. Serial determinations of LH, FSH, and estradiol done prospectively on five consecutive days in 19 of the 90 patients did not give any further information in differentiating among the four groups.