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PubMed1985

[A combined radioimmunoassay method for the determination of 17 alpha-hydroxypregnenolone, 17 alpha-hydroxypregnenolone sulfate and 17 alpha-hydroxyprogesterone in human blood].

It is well recognized that in the fetal adrenal cortex, 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) activity is lower and in fetal tissues, steroid sulfokinase (SK) is higher than in the adult. In order to clarify a part of the development processes of the adrenal cortex or steroidgenesis in humans, a combined radioimmunoassay (RIA) method to estimate serum 17 alpha-hydroxypregnenolone (17-OH-delta 5 P), 17 alpha-hydroxypregnenolone sulfate (17-OH-delta 5 P-S) and 17 alpha-hydroxyprogesterone (17-OH-delta 4P) was devised. The method consisted of the following procedures: 1) diethyl ether extraction and chromatographic separation of unconjugated steroids (17-OH-delta 5P and 17-OH-delta 4P), 2) enzymatic hydrolysis of 17-OH-delta 5P-S using the residue of diethyl ether extraction for a material, 3) diethyl ether extraction and chromatographic purification of hydrolyzed 17-OH-delta 5P-S, and 4) RIAs for 17-OH-delta 5-P to estimate 17-OH-delta 5P and 17-OH-delta 5P-S concentration, and for 17-OH-delta 4P. Extracted 17-OH-delta 5P was well separated from 17-OH-delta 4P by Sephadex LH-20 microcolumn chromatography, using a benzene/methanol = 95/5 (v/v) solvent as a mobile phase. Several procedures for hydrolysis or solvolysis of 17-OH-delta 5P-S were compared using available tritiated delta 5-3 beta-hydroxysteroids including dehydroepiandrosterone (DHA), DHA sulfate (DHA-S) and 17-OH-delta 5P, and it was found that the most suitable method was an enzymatic hydrolysis by arylsulfatase from Helix Pomatia in an appropriate condition in which the percent hydrolysis was 92.9 +/- 1.2 (mean +/- SEM)%. The final percent recoveries were 88.7 +/- 1.2% in 17-OH-delta 4P, 90.7 +/- 1.4% in 17-OH-delta 5P and 78.1 +/- 2.1% in 17-OH-delta 5P-S, respectively. A suitable antiserum and its final dilution titer for RIA of 17-OH-delta 5P (hydrolyzed 17-OH-delta 5P-S also) was 1:12,000 dilution of anti-17-OH-delta 5P-3-succinate-BSA serum. An anti-7-oxo-17-OH-delta 5P-7-carboxymethyloxime-BSA serum was considered to be unsuitable for the measurement of hydrolyzed 17-OH-delta 5-P-S, presumably because of a significant cross-reactivity with a large amount of unknown steroid sulfates simultaneously hydrolyzed.(ABSTRACT TRUNCATED AT 400 WORDS)

17-alpha-Hydroxypregnenolone↗
PubMed1986

[Neonatal screening for congenital adrenal hyperplasia due to 21-hydroxylase deficiency. 1. Enzyme immunoassay of dried blood 17 alpha-hydroxyprogesterone and its application to neonatal screening for congenital adrenal hyperplasia].

An enzyme immunoassay for measuring 17 alpha-hydroxyprogesterone (17-OHP) in dried blood collected on filter paper has been developed. The method is easy and rapid and has specificity, accuracy and precision. 17-OHP values of neonates with congenital adrenal hyperplasia (CAH, 40 ng/ml) were extremely high compared with normal neonates (1.1 +/- 0.7 ng/ml). There was a negative correlation between the 17-OHP value and birth weight. The method has been applied to neonatal screening for CAH due to 21-hydroxylase deficiency. During 38 months, 67,392 neonates were screened. The recall rate and the medical evaluation rate were 1.16% and 0.09%, respectively. A third of recalled neonates were low birth weight infants. 5 neonates were proven to have CAH, and its incidence was 1:13,478. The present study demonstrates the feasibility of a neonatal screening for CAH and indicates that the frequency of CAH may be greater than previously reported by case assessment method in Japan.

17-alpha-Hydroxyprogesterone↗
PubMed1991

[Determination of urinary 17 alpha-hydroxyprogesterone excretion using ELISA--evaluation of normal subjects and patients with congenital adrenal hyperplasia due to 21-hydroxylase deficiency].

The utility of urinary 17 alpha-hydroxyprogesterone (U-17-OHP) in the diagnosis and management of congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency (21-OHD) was evaluated in 16 patients with 21-OHD. The normal values for U-17-OHP in relation to age and other physiological conditions were investigated in 96 normal subjects and 7 low birth weight infants. Levels of U-17-OHP, serum 17-OHP (S-17-OHP) and urinary free cortisol (U-FC) were simultaneously determined using enzyme-linked immunosolvent assay (ELISA) combined with fractionation by high performance liquid chromatography (HPLC). Pregnanetriol (PT) levels in the same urine specimens were determined using glass capillary gas chromatography (GC). The results were as follows: 1) Normal subjects and low birth weight infants. A significant correlation between U-17-OHP excretions corrected for body surface area (BSA) in 2-h urine specimens and S-17-OHP concentrations at the midpoint of the urine sampling period was observed in normal subjects (r = 0.768, p less than 0.01). Circadian U-17-OHP excretion in 6 adult males was synchronous with that of U-FC. Age-related changes in actual U-17-OHP excretions (Mean +/- SD ng/day) were as follows: neonates: 31.9 +/- 10.3, children aged 2 to 4 years old: 29.1 +/- 14.5, 5 to 8 years old: 68.6 +/- 29.9, 9 to 11 years old: 151.3 +/- 50.0, 12 to 15 years old: 222.7 +/- 82.0, adult males: 400.1 +/- 62.5, adult females (luteal phase): 339.1 +/- 109.7, and adult females (follicular phase): 185.6 +/- 72.3, respectively. Correlation between U-17-OHP and PT excretions in 24-h urine specimens from normal subjects greater than 2 years of age was highly significant (r = 0.871, p less than 0.01). Although U-17-OHP values were measurable in neonates, those of PT were not detectable by GC in the same specimens because of low conversion of 17-OHP to PT. 2) Cases of 21-OHD. In 4 cases of 21-OHD diagnosed in the neonatal period (aged 11 to 15 days, all were of the salt-losing type), the U-17-OHP concentration in a single urine specimen was significantly higher than that of age-matched controls, whereas the PT concentration in one case was low and therefore had no diagnostic value. In 12 patients with 21-OHD receiving suppressive therapy, correlation between S-17-OHP concentrations and 24-h U-17-OHP excretions corrected for BSA was significant (r = 0.847, p less than 0.01).(ABSTRACT TRUNCATED AT 400 WORDS)

17-alpha-Hydroxyprogesterone↗
PubMed2005

Hydroxyprogesterone caproate and progesterone increase tumor necrosis factor-alpha production in lipopolysaccharide stimulated whole blood from non-pregnant women.

OBJECTIVE: Hydroxyprogesterone caproate (17-P) and progesterone have been shown to decrease the rate of preterm birth in high-risk pregnant women, but the mechanism of action is unknown. We hypothesized that 17-P or progesterone would reduce production of the pro-inflammatory cytokine, tumor necrosis factor-alpha (TNF-alpha), after lipopolysaccharide (LPS) stimulation of whole blood. STUDY DESIGN: Whole blood collected from 10 non-pregnant women in the follicular phase was treated with LPS (1000 ng/mL) alone or with either 17-P or progesterone (125 ng/mL and 1250 ng/mL) and LPS. Supernatant collected after 24 h was tested for TNF-alpha by enzyme immunoassay. Results were compared using the Wilcoxon rank sum test. RESULTS: Whole blood treated with 17-P or progesterone in addition to LPS produced significantly higher TNF-alpha concentrations than blood treated with LPS alone. CONCLUSION: 17-P and progesterone appear to have a pro-inflammatory effect during LPS stimulation of blood from non-pregnant women in vitro. Our data suggest that the reduction in pre-term birth in women treated with progesterone is not mediated through an anti-inflammatory effect on peripheral blood cells.

17 alpha-Hydroxyprogesterone Caproate↗
PubMed1986

Failure of GnRH analogue to inhibit serum concentrations of testosterone and 17 alpha-hydroxyprogesterone in hCG-substituted hypogonadotropic hypogonadism.

Gonadotropin-releasing hormone analogues (GnRH-A) induce inhibition of testicular function and reduction of serum testosterone (T) in man, but the mechanism involved is still debatable. To elucidate it we studied six patients with hypogonadotropic hypogonadism (HH) in chronic substitution with hCG for correction of androgen deficiency symptoms, and evaluated the effect of addition of GnRH-A to the hCG therapy on plasma levels of T and 17 alpha-hydroxyprogesterone (17 OHP). All patients were treated with 1000 U of hCG in every 3rd day for 24 weeks. After 8 weeks of this regimen, GnRH-A, Buserelin (D-Ser-TBU-EA-LHRH), 200 micrograms per day sc, was added and given for 8 weeks. After cessation of analogue administration patients were followed for 8 further weeks. The levels of the two steroids did not differ markedly in the pre- and post-GnRH-A period. GnRH-A given for two months did not lower T or 17 OHP levels as in eugonadal men after similar treatment. The median T concentrations during GnRH-A tended to be increased, with plasma values higher (P less than 0.025) than the peak values observed during hCG alone. Since administration of Buserelin did not inhibit hCG-sustained steroid levels in these HH patients, it is conceivable that GnRH-A may have lacked a direct inhibitory gonadal effect in such experimental conditions.

17-alpha-Hydroxyprogesterone↗
PubMed1985

Photoperiod-induced changes in the testicular metabolism of [4-14C]17 alpha-hydroxyprogesterone in the bank vole (Clethrionomys glareolus).

Minces of the testes of bank voles, born and reared in a long (18L:6D) photoperiod until weaning (18-22 days of age) and subjected thereafter to a short (6L:18D, Group S) or a long (18L:6D, Group L) photoperiod for 6-9 weeks, were incubated with [4-14C]17 alpha-hydroxyprogesterone in the presence of cofactors (NADP/NADPH, 1.3 mmol/1) for 1 h at 37 degrees C. The radioactive metabolites were characterized and identified by thin-layer chromatography with derivative formation and chromatography to constant specific activity and isotope ratio. In Group L virtually all of the substrate was utilized and it was readily converted to androgens (48% of the radioactivity recovered) such as androstenedione and testosterone. The only pregnane metabolite identified was 17 alpha-hydroxy,20 alpha-dihydroxyprogesterone (43.3%). In Group S there was a decreased production of 17 alpha-hydroxy,20 alpha-dihydroprogesterone and androgens (25.4% and 10.4% respectively) and a substantial portion of the substrate was not metabolized (38.8%). The main androgen metabolites identified, androst-4-ene-3 beta,17 beta-diol and 5 alpha-androstane-3,17-dione are hormonally quite inert steroids. No androstenedione or testosterone was found. The results indicate that exposure to short photoperiod induces a decrease in the testicular C17-C20 lyase and 20 alpha-hydroxysteroid dehydrogenase.

17-alpha-Hydroxyprogesterone↗
PubMed1985

Serum lipids and lipoproteins in patients with endometrial carcinoma receiving adjuvant treatment with hydroxyprogesterone caproate.

Serum levels of triglycerides, total cholesterol, and HDL-cholesterol were determined in 57 patients who were undergoing treatment for stage I endometrial carcinoma. The patients belonged to a clinical trial where group A (control) was treated with surgery plus intravaginal irradiation, whereas group B in addition was treated with hydroxyprogesterone caproate (5000 mg i.m. as a loading dose followed by 1000 mg every 2 weeks for one year). In group B patients followed during the first 13 weeks of treatment, the level of serum triglycerides remained stable, whereas the level of total cholesterol and HDL-cholesterol increased significantly. This increase could not, however, have been caused by the progestogen treatment, as similar changes were seen in group A patients followed for the same period of time. Long-term effects were looked for in patient groups examined 3-12 months after the start of treatment and in groups examined 3-6 months after the hormone therapy was stopped. In neither group could any significant difference in cholesterol or HDL-cholesterol be found. It is concluded that this type of progestogen treatment causes no significant change in the levels of triglycerides, cholesterol, and HDL-cholesterol.

17 alpha-Hydroxyprogesterone Caproate↗
PubMed1985

Intramuscular administration of hydroxyprogesterone caproate in patients with endometrial carcinoma. Pharmacokinetics and effects on adrenal function.

A radio-immunoassay for the determination of the serum concentration of hydroxyprogesterone caproate (HPC) was established. After a single intramuscular injection of 1000 mg, the mean serum level reached its maximum (44-81 nmol/l) after 2-7 days. Patients on long-term adjuvant HPC treatment (consisting of 1000 mg daily for 5 days followed by 1000 mg every 2 weeks) presented peak hormone levels 2 weeks after commencing treatment. After a drop at 5 weeks, the mean serum level slowly increased again to 130 nmol/l after 25 weeks of treatment. Patients being treated with weekly injections had significantly higher serum levels than those treated every 2 weeks. Considerable inter-individual differences were observed. The serum concentrations of HPC measured in this study compare favorably with those previously found in patients treated with medroxyprogesterone acetate. The patients on adjuvant HPC showed no significant change in the levels of cortisol, dehydroepiandrosterone sulphate, androstenedione, or estrone during the first 25 weeks of treatment.

17 alpha-Hydroxyprogesterone Caproate↗
PubMed1988

Development of a direct [125I]radioimmunoassay for serum levels of 17-hydroxyprogesterone.

We describe a direct, rapid, sensitive and highly specific radioimmunoassay for determining serum levels of 17-hydroxyprogesterone. It is based on the use of highly specific sheep antiserum, 125I-labelled tracer, dextran-coated charcoal to separate the antibody bound and free fractions, and sodium salicylate to eliminate interference from endogenous binding proteins in serum. Intra- and interassay coefficients of variation are less than 8% and recovery is satisfactory. Sensitivity is 3.5 fmol per assay tube (0.14 nmol/l). Results correlate closely with those of an established technique using 3H-labelled steroid after initial solvent extraction and column chromatography of samples (y = 1.02x - 0.16; r = 0.998). The values found for serum from normal adult subjects ranged from 1.0 to 12.1 nmol/l while those from treated and untreated patients with congenital adrenal hyperplasia due to 21-hydroxylase deficiency were 1.4-18.4 and 27.3-650 nmol/l, respectively.

17-alpha-Hydroxyprogesterone↗
PubMed1987

Trial of 17-hydroxyprogesterone caproate (Proluton Depot) in women with long-standing infertility; failure of estrogen positive feedback the following cycle.

In open and double-blind studies 40 women with long-standing unexplained infertility were investigated and treated with 17 alpha-hydroxyprogesterone caproate (17HPC, Proluton Depot). In the open study, 16 women with a high index of suspicious abortions were given 500 mg 17HPC imtramuscularly (i.m.) weekly for 6-16 weeks. Six of the women received the drug from a time preceding the expected date of a period; 2 of these conceived that cycle and their pregnancies continued to term, while 4 were not pregnant. Ten women (9 with definite previous abortions) were given 17HPC when they suspected (correctly) that they were pregnant. Their pregnancies continued to term in all but 1, who had a premature delivery (still-birth) at 34 weeks. In the double-blind study 24 women were given injections of 17HPC or placebo i.m. at weekly intervals, from about 4 days before the expected period (day -4), provided that the level of progesterone (Prog) (on days -9 to -7) was greater than 10 nmol/l. After placebo no delay in menstruation or disruption of the succeeding cycle was observed. In 14 of 16 cycles in 14 women given 500 mg 17HPC the withdrawal period was delayed by a few days, and then followed by highly erratic bleeding over the next 1-3 months. The dose was therefore reduced to 250 mg 17HPC but the same problem resulted in 8 of 29 cycles in 16 women (including the above studied in later cycles). Regular cycles were eventually restored in all cases but in 3 this necessitated treatment with the contraceptive pill (Microgynon). In most of the post-treatment cycles there was a progressive and prolonged estradiol (E2) rise, which was not preceded by changes either in serum FSH concentration or in the LH/FSH ratio nor associated with the expected positive feedback rise in LH. We conclude that 17HPC disrupts the following cycle, probably by allowing follicular development while interfering with positive LH feedback. None of the patients of the double-blind study had conceived (as evidenced by undetectable hCG levels). Our study confirmed that this progestogen exerts no direct luteolytic effect. However, in order to establish the efficacy or otherwise of 17HPC given before the end of the cycle, women should be selected with a very high index of suspicion of recurrent early implantation failure or abortion.

17 alpha-Hydroxyprogesterone Caproate↗
PubMed1992

Simultaneous quadruple-label fluorometric immunoassay of thyroid-stimulating hormone, 17 alpha-hydroxyprogesterone, immunoreactive trypsin, and creatine kinase MM isoenzyme in dried blood spots.

We describe a quadruple-label fluorometric immunoassay for simultaneously measuring four analytes: thyroid-stimulating hormone (TSH), 17 alpha-hydroxyprogesterone (17 alpha-OHP), immunoreactive trypsin (IRT), and creatine kinase MM (CK-MM). The assay is based on immunoreagents labeled with four different lanthanide ions (Eu3+, Tb3+, Sm3+, and Dy3+), on dissociative fluorescence enhancement applying the principle of co-fluorescence, and on time-resolved fluorometry. The monoclonal anti-alpha-TSH and anti-IRT antibodies and the polyclonal anti-CK-MM antibody were labeled with Eu3+, Sm3+, and Dy3+, respectively; 17 alpha-OHP was labeled with Tb3+. The assay was performed in microtitration strip wells coated with a mixture of monoclonal antibodies against beta-TSH, IRT, and CK-MM and a polyclonal goat anti-rabbit IgG for capture of the rabbit anti-17 alpha-OHP antibodies. After completion of the immunoreactions, the bound fractions of the lanthanides were dissociated into the co-fluorescence enhancement solution, creating highly fluorescent chelates. The four lanthanide-specific signals were subsequently measured in a time-resolved fluorometer. The detection limits of the assay were 0.1 mIU/L for TSH, 2 nmol/L for 17 alpha-OHP, 2 micrograms/L for IRT, and 4 U/L for CK-MM.

17-alpha-Hydroxyprogesterone↗
PubMed1992

Detection of testosterone administration by increased ratio between serum concentrations of testosterone and 17 alpha-hydroxyprogesterone.

An increased ratio between urinary testosterone (T) and epitestosterone (epiT) has been accepted by the International Olympic Committee as a marker for T doping. However, in a few subjects, we and others have observed constantly above-normal urinary T/epiT ratios that are unlikely to be related to exogenous T administration. To find a better test for T doping, we studied several serum and urinary androgens and androgen precursors, estrogens, and luteinizing hormone (LH) in seven healthy volunteers for 35 days after an intramuscular injection of 250 mg of testosterone enanthate. Among urinary analyses, only the T/epiT ratio was a suitable marker of T doping; of the serum assays, 17 alpha-hydroxyprogesterone (17OHP), T/17OHP ratio, LH, and T/LH ratio were fair to good markers of T doping. The serum T/17OHP ratio was the best marker of those tested, with all seven subjects having above-normal values for this in the first 3 days of the observation period. No other marker showed abnormal values in all subjects at any time. Moreover, the T/17OHP ratio was affected by neither diurnal variation nor physical stress. The value of this marker for T doping was further supported by the finding of normal T/17OHP ratios in a subject with increased urinary T/epiT ratios caused by an abnormally low testicular epiT production, probably related to genetic factors.

17-alpha-Hydroxyprogesterone↗
PubMed1992

Identification of the steroids in neonatal plasma that interfere with 17 alpha-hydroxyprogesterone radioimmunoassays.

Neonatal plasma contains interferents that increase the apparent 17 alpha-hydroxyprogesterone (17-OHP) content measured by direct (no-extraction) radioimmunoassay. We fractionated extracts from neonatal plasma pools by liquid chromatography with a Sephadex LH-20 column and measured 17-OHP immunoreactivity by a direct test kit. We found immunoreactivity in the free steroid and glucuronide fraction and also in the monosulfate fraction. We analyzed these two fractions by reversed-phase high-performance liquid chromatography (HPLC) and HPLC-mass spectrometry. We collected fractions and assayed for 17-OHP immunoreactivity. The HPLC fractions containing the interfering steroid monosulfates were analyzed by ion-spray mass spectrometry and, after solvolysis, by gas chromatography-mass spectrometry. Several monosulfates were identified, including those of 17 alpha-hydroxy-pregnenolone, 16 alpha-hydroxypregnenolone, pregnenolone, and several pregnenetriols. 17 alpha-Hydroxypregnenolone sulfate was the most significant interferent. Other commercially available 17-OHP assays showed similar interference when used without an extraction step. Kit manufacturers should select antibodies and protocols to minimize cross-reaction with sulfates, especially 17 alpha-hydroxypregnenolone sulfate.

17-alpha-Hydroxyprogesterone↗
PubMed2005

17 alpha-hydroxyprogesterone caproate (17P) usage in a Medicaid managed care plan and reduction in neonatal intensive care unit days.

PURPOSE: To evaluate whether providing 17 alpha-hydroxyprogesterone caproate (17P) to high-risk pregnant women who have a history of preterm delivery in a Medicaid managed care population reduces the rate of neonatal intensive care unit (NICU) admissions, NICU length of stay, and associated costs. DESIGN: A 2004-2005 longitudinal review of birth outcomes in 24 pregnant women with a history of preterm delivery who were treated with 17P versus a control group. METHODOLOGY: Intervention included offering 17P as a benefit to pregnant women who had a history of preterm labor and delivery and who were deemed to be appropriate candidates for this treatment by their physicians. An educational program about 17P was developed that was aimed at physicians, their office staff, and plan members. A process of early identification of potential 17P candidates was also implemented. PRINCIPAL FINDINGS: NICU admission rates decreased to 14.3 percent in the control group and 8.3 percent in the 17P group. NICU length of stay decreased significantly from 231 days in the control group to 149 days in the 17P group. Overall costs for the control group were dollar 568,462 versus dollar 165,487 in the treatment group--a significant savings of dollar 402,975. CONCLUSION: Offering 17P as a benefit to pregnant women enrollees with a history of preterm delivery can decrease NICU days significantly for a Medicaid managed care plan.

17 alpha-Hydroxyprogesterone Caproate↗
PubMed1991

Age of appearance of circadian rhythm in blood 17-hydroxyprogesterone in 21-hydroxylase deficiency.

Sixty-one daily profiles of blood-spot 17-hydroxyprogesterone derived from patients with congenital adrenal hyperplasia due to 21-hydroxylase deficiency were analysed to determine the age of appearance of circadian rhythm in hypothalamo-pituitary-adrenal activity. The results revealed that typical circadian adrenocortical rhythm is established at approximately three months after birth. In conclusion, the maturation of supra-hypothalamic brain structure proceeds normally also in humans with constant low level of cortisol in blood.

17-alpha-Hydroxyprogesterone↗
PubMedDate not supplied

Influence of progesterone and 17 alpha hydroxyprogesterone upon the BANA-hydrolytic activity of human sperm.

Our knowledge about temporal relationships between reproductive processes and defined changes in the plasmatic concentrations of 17 beta-estradiol, Follicle-stimulating hormone (FSH), Luteinizing hormone (LH) and progesterone (P) is still incomplete. Is known that in periovulatory phase the chance of fertilization increases to its maximum. The results obtained using different concentration of P have shown that at high concentration a fast liberation or/and exposure of the BANA-hydrolytic (B-H) activity is present. However, with low concentrations of P the enzymatic activity keeps a relation with the exposure time. In similar experimental conditions and using 17 alpha-hydroxyprogesterone it has been no change in the B-H activity. The results obtained in the present study suggest that the P possibly acts upon the sperm stimulating its hydrolytic activity, allowing its penetration through the zona pellucida of the ovum.

17-alpha-Hydroxyprogesterone↗
PubMedDate not supplied

17-Hydroxyprogesterone level as a marker for corpus luteum function in aborters versus nonaborters.

Weekly serum levels of 17-hydroxyprogesterone (17-OHP) levels and serum progesterone (P) levels were measured in 378 pregnant women. Normal levels of 17-OHP were established in women taking ovulation-inducing drugs (OVID) versus those conceiving on normal cycles. Weekly levels of 17-OHP were measured in women who aborted and were compared with the established norms. The levels of 17-OHP in aborters were lower than in normals for their specific category established in nonaborters. These results suggest that a failing corpus luteum may contribute to some abortions. Careful attention to 17-OHP levels dropping below normal during the first trimester may alert the clinician to the need to increase progesterone exogenously and possibly prevent an abortion.

17-alpha-Hydroxyprogesterone↗
PubMed1990

Direct solid-phase time-resolved fluoroimmunoassay of 17 alpha-hydroxyprogesterone in serum and dried blood spots on filter paper.

We describe a direct, solid-phase time-resolved fluoroimmunoassay (TRFIA) for measuring 17 alpha-hydroxyprogesterone (17OHP) in serum and blood spots on filter paper. We used 17OHP-3-carboxymethyloxime (17OHP3CMO) coupled to polylysine as the label, which enabled incorporation of up to 34 atoms of europium per molecule of 17OHP, for a very high specific activity. The assay is based on competition between labeled 17OHP3CMO and 17OHP in blood specimens for polyclonal rabbit anti-17OHP antibodies. The antibody-label complex is separated by binding to anti-rabbit antibodies coated onto microtiter strips. The assay buffer contains danazol to displace 17OHP from steroid-binding proteins in serum. For serum samples, the assay is accomplished in 1 h of incubation at room temperature. The blood spot assay with filter paper discs involves incubation overnight at 4 degrees C. Results for both types of specimens from the same subjects correlated well. The lowest measurable concentrations of 17OHP (nmol/L) were 0.10 (3 SD) and 0.75 (3 SD) for serum and dried blood on filter paper, respectively. Intra- and interassay CVs were about 5-15% for both types of samples.

17-alpha-Hydroxyprogesterone↗