Sergio C. Stone

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

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.

Inhibition of ovulation with cyclic use of progestogen-impregnated intravaginal devices

The efficacy acceptability and reversibility of contraception with silicone vaginal rings impregnated with medroxyprogesterone acetate were studied using 24 female subjects (age 24-39) over a 10 month period. For all these subjects there was presumptive evidence of ovulation in 2 control cycles. During each treatment cycle new devices were inserted either on Day 5 or Day 10 and removed on Day 26. The women were studied for 2 posttreatment cycles. During treatment ovulation was inhibited as evidenced by failure to detect a rise of serum progesterone in the luteal phase of the cycle. Ovulation as determined by this parameter usually resumed in the first cycle after treatment. There was minimal breakthrough bleeding. Small vaginal erosions in 4 subjects healed spontaneously following removal of the devices. The subjects and their husbands stated that the device did not cause discomfort during coitus. After initial fitting by a physician the women removed and reinserted the rings without difficulty. This technique offers promise as a method of contraception.((authors modified))

Serum estradiol in women ingesting combination oral contraceptive steroids

n A study of the effect of combination oral contraceptives upon endogenous ovarian estrogen production was done by measuring serum estradiol levels in 3 groups of women. The first group of 3 women gave daily serum samples during 1 control and 2 treated cycles (2 mg. norethindrone and 100 mu-g mestranol received daily for 20 days), and levels of follicle-stimulating hormone, luteinizing hormone, estradiol and progesterone were measured. A second group (6) taking oral contraceptives for at least a year were measured daily during 1 cycle for estradiol only. Each of 97 women in the third group, taking the pill from 1 to 6 years, gave a single blood sample taken at random during the cycle, to be assayed for estradiol. Estradiol levels in women receiving oral contraceptives were low, usually in the range of 20-30 pg./ml., similar to those found in the early follicular phase in ovulatory cycles, and significantly higher than levels found in postmenopausal women. Exogenous estrogen in the pill, together with this level of endogenous estradiol, should be sufficient to prevent any harmful effects associated with estrogen deficiency. The finding that estradiol levels remain low in the first cycle of therapy is consistent with previous studies which indicate that one of the mechanisms of action of hormonal contraceptives is a direct effect upon the ovary.n

A modified technique for the assay of progesterone in blood using celite column chromatography

Abstract A modified technique for the assay of progesterone in blood is presented. The procedure consists of three main steps: 1) ether extraction, 2) rapid and complete separation of progesterone from more polar steroids by the use of celite column chromatography, and 3) radioassay of progesterone by competitive protein binding. This assay method has a sensitivity of 0.1 ng and excellent specificity with low values for blank samples. The precision of this assay technique is comparable with other reported methods. This method offers two main advantages in comparison with previously described procedures. First, the celite column chromatography provides a rapid technique of separation of progesterone from other cross reacting steroids with a recovery of greater than 75 per cent. Secondly, the more polar steroids can be eluted subsequently and assayed separately. Thus, the assay of other steroids can be easily performed using the same serum extract.

A technique for the assay of 17-hydroxyprogesterone (17-hydroxy-4-pregnene-3,20-dione) in serum using celite column chromatography and competitive protein binding

Abstract An improved method for the assay of 17-hydroxyprogesterone (17-hydroxy-4-pregnene-3,20-dione) in blood is presented. The procedure utilizes a Celite column which is more rapid and yields lower blanks than thin layer chromatography. It also allows the separation and assay of other steroids in the same extracted sample. The sensitivity and precision reported are comparable to other methods. The mean values obtained for men were 1.0 ng/ml, for women in the follicular phase 0.3 ng/ml and in the luteal phase 1.3 ng/ml. These values agree closely with other reports.

Cross-reaction of contraceptive steroids in competitive binding assays of progesterone and estradiol

n Cross-reaction of oral contraceptives (OCs) in competitive binding assays of progesterone and estradiol was investigated. The compounds tested were chlormadinone acetate, diethylstilbestrol, dimethisterone, dydrogesterone, ethinyl estradiol, ethynodiol diacetate, megestrol acetate, mestranol, norethynodrel, norethindrone, d-norgestrel, medroxyprogesterone acetate, quinestrol, and quingestanol acetate. Only ethinyl estradiol had a cross-reaction of more than 1% (2%) in the estradiol radioimmunoassay and dydrogesterone (1.4%) and d-norgestrel (4%) were the only compounds that showed a cross-reaction more than 1% in the progesterone competitive protein-binding assay. These results indicate that the steroids tested, in doses currently being used in OCs, will not markedly interfere with the assay of endogenous estradiol and progesterone.n