Prospective assessment of early fetal loss using an immunoenzymometric screening assay for detection of urinary human chorionic gonadotropin.

OBJECTIVE: To develop an economical, nonradiometric immunoenzymometric assay (IEMA) for the detection of urinary human chorionic gonadotropin (hCG) in studies of early fetal loss. To be effective, the IEMA must have a sensitivity equal to the standard immunoradiometric assay (IRMA) and sufficient specificity to eliminate the need for screening most nonconceptive cycles with the expensive and labor-intensive IRMA. DESIGN: Two different assays were used to measure hCG in daily early morning urine samples from potential conceptive cycles. SETTING: Women undergoing donor artificial insemination (AI) were evaluated in a prospective study. PATIENTS, PARTICIPANTS: Ninety-two women volunteers were selected on the basis of apparent normal reproductive health. INTERVENTIONS: Artificial insemination with nonfrozen donor semen was performed by cervical cup twice each menstrual cycle at 48-hour intervals, and daily urine samples were self-collected throughout the menstrual cycle. MAIN OUTCOME MEASURES: An IEMA was developed to detect urinary hCG using the same antibodies as in the standard IRMA; a study was designed to determine whether this nonradiometric assay could successfully detect the early fetal loss that was detected by the IRMA. RESULTS: Of 224 menstrual cycles analyzed by both assays, a total of six early fetal losses were detected by the IRMA. When the tentative screening rule was set to allow all six of these losses and 95% of future losses to be detected by the IEMA, an additional 34 false-positive results were detected by the IEMA. The specificity of the IEMA with this rule was calculated to be 84%. CONCLUSION: An IEMA based on the same antibodies used for the standard IRMA can serve as an efficient screening assay for the detection of early fetal loss. When the IEMA is used in this manner, nearly 80% of screened menstrual cycles can be eliminated without further testing by the IRMA.     

A semiquantitative human chorionic gonadotropin assay for the detection of ectopic pregnancy

Human chorionic gonadotropin (hCG) was measured in 117 serum samples with known quantities of hCG after a dilutional modification of a reliable, simple, and inexpensive qualitative assay for hCG. The modification yielded a semiquantitative assay for hCG with a sensitivity of 5000 mIU/mL. At hCG concentrations below 4000 mIU/mL, the assay had no false-negative or false-positive results; above 6500 mIU/mL, there were also no false-negative or false-positive results. In the range of 4000-6500 mIU/mL, the clinical false-positive rate was 28%. Using the described dilutional modification of this qualitative hCG assay, the test is semiquantitative, and is useful in selecting the appropriate time to perform ultrasound and laparoscopy in women suspected of having an ectopic pregnancy.     

The effect of different human chorionic gonadotropin assay sensitivity on screening for ectopic pregnancy

The distribution of human chorionic gonadotropin levels in 184 patients with ectopic pregnancy is examined. The impact of changing the sensitivity of human chorionic gonadotropin testing on the incidence of false negative results is discussed. Pregnancy blood tests with a detection threshold of 200 mIU/ml were associated with an 11.9% incidence of false negative results.     

Characterization of human chorionic gonadotropin peptide variants with a radio-receptor assay using recombinant human luteinizing hormone/chorionic gonadotropin receptors

There are potential interactions between various human chorionic gonadotropin (hCG) isoforms at the level of luteinizing hormone/chorionic gonadotropin (LH/CG) receptor. The objective of this study was to characterize the receptor-binding activities of the primary peptide variants of hCG including intact hCG, free beta subunit, beta-core fragment and nicked hCG, and to test the effects of these hCG variants on the binding of intact hCG. A radio-receptor assay based on cell membranes expressing recombinant human LH/CG receptors was validated and used in this study to avoid species differences in the receptor-binding specificity. The results showed that none of the hCG variants that we studied had sufficient binding affinity to compete with binding of intact hCG, nor were they able to antagonize the binding of intact hCG. These results suggest that hCG variants with either abbreviated polypeptide structures or incomplete peptide linkage are products or metabolites which do not have the tropic biological activity of the whole hormone, the intact heterodimeric hCG.     

Reverse passive hemagglutination assay for human chorionic gonadotropin in urine using monoclonal antibody

Aiming to find a urinary hCG immuno-assay which is specific, sensitive and easy to perform, a reverse passive hemagglutination reaction was studied by using sheep red blood cells (SRBC) coupled with monoclonal antibodies (Mab) to hCG. Three Mabs (5D4, 6E4, 2F8) with different specialty were used for the study. Mab 5D4 reacted to hCG, hCG-beta, and LH but not to hCG-alpha. Mab 6E4 reacted to hCG, hCG-alpha and LH, but not to hCG-beta. Mab 2F8 reacted to hCG but not to hCG-alpha, hCG-beta, or LH. All three Mabs were IgG1. SRBC were treated with glutaraldehyde and then with tannic acid. These treated SRBC were coupled with IgG(2mg/ml) of each anti hCG-Mab. For assays, 30 microliters of 1:1 mixtures of two different Mab-coupled SRBC and 30 microliters of standard hCG or urine samples were mixed in wells of microtiter plates and reacted for 60 min at room temperature. Among three different combinations, the couple 5D4-SRBC and 2F8-SRBC were most sensitive and specific for hCG assays and the minimum amount of hCG and LH detected in this combination assays was 12.5 mIU/ml and 800 mIU/ml, respectively. Some clinical data obtained by applying this assay were presented.     

A single chorionic gonadotropin assay for maternal serum screening for Down's syndrome

A simple enzyme immunoassay measuring human chorionic gonadotropin in undiluted maternal serum has been developed in order to be used as a prenatal screening test for Down's syndrome. A retrospective study of maternal serum sampled during pregnancies associated with trisomy 21 shows that with a 5% amniocentesis rate determined on a single test, the detection rate of trisomy 21 would be around two-thirds of the affected pregnancies. A prospective study of 9040 pregnant women under 38 years has confirmed the usefulness of the assay.     

Prospective intervention trial of a screening protocol to identify fetal trisomy 18 using maternal serum alpha-fetoprotein, unconjugated oestriol, and human chorionic gonadotropin.

Two prenatal centres in New England, routinely using a screening protocol for fetal Down syndrome that included maternal serum alpha-fetoprotein (AFP), unconjugated oestriol (uE3), and human chorionic gonadotropin (hCG) measurements in combination with maternal age, adopted a separate screening protocol for trisomy 18. That protocol identified a pregnancy as being at high risk when AFP, uE3, and hCG measurements all fell at or below specified cut-offs (0.75, 0.60, and 0.55 multiples of the median, respectively), regardless of maternal age. Among the first 19,491 women screened, 98 (0.5 per cent) were found to have values which placed them in the high-risk category. Four of these women were subsequently found not to be pregnant. In two others, samples from non-pregnant individuals were found to have been incorrectly submitted for analysis in place of the samples from the pregnant women. All of the remaining 92 women were counselled and offered amniocentesis and fetal karyotyping. Eighty-eight (96 per cent) accepted. Karyotypes or birth outcomes were available on all 92 pregnancies. Six cases of trisomy 18 and one case of Turner syndrome were identified by karyotype. One case of trisomy 18 was identified for every 14 unaffected pregnancies offered amniocentesis. In the present prospective study, an estimated 85 per cent of the cases of trisomy 18 were identified. However, given the small number of cases (six), the 95 per cent confidence interval for the detection rate is broad (40-95 per cent).     

Prospective evaluation of free beta-subunit of human chorionic gonadotropin and dimeric inhibin A for aneuploidy detection.

OBJECTIVE: Our goal was to prospectively evaluate the use of the free beta-subunit of human chorionic gonadotropin and dimeric inhibin A for the detection of fetal Down syndrome and other aneuploidies. STUDY DESIGN: Women who had a second-trimester multiple-marker screening test (alpha-fetoprotein, unconjugated estriol, human chorionic gonadotropin) and genetic amniocentesis from August 1996 to August 1998 were included. Serum was also analyzed for inhibin and the free beta-subunit of human chorionic gonadotropin. Detection and false-positive rates for 4 analyte combinations at 5 different screening risk cutoff points for Down syndrome were determined and compared. RESULTS: We evaluated 1256 patients, including 23 with aneuploidy (13 with Down syndrome, 10 others). The maternal age was 35.9 +/- 4.6 years (mean +/- SD). At the optimal risk cutoff point for Down syndrome detection (1:190; false-positive rate, 19%), the multiple-marker screening test plus inhibin was superior, detecting 85% of Down syndrome cases, in comparison with 69% when the multiple-marker screening test alone was used and 62% when the other 2 combinations were used. The multiple-marker screening test plus inhibin also detected 60% of the other aneuploidies. CONCLUSIONS: When evaluated prospectively in a high-risk population, the multiple-marker screening test plus inhibin was superior to the traditional multiple-marker screening test and 2 other analyte combinations, with a lower false-positive rate and increased detection of all aneuploidies in a high-risk population.     

Use of an immunoradiometric assay and a radioimmunoassay for the detection of free beta-human chorionic gonadotropin

An immunoradiometric assay and a radioimmunoassay (RIA) were used to quantitate human chorionic gonadotropin (hCG) in the sera of ten pregnant women at term and of six women with gestational trophoblastic neoplasia. The two techniques show good correlation (Pearson correlation coefficient .96) in the assay of pregnancy serum. Because only the RIA, and not the immunoradiometric assay, measures the free beta-subunit of hCG, a comparison of the results obtained by the two immunoassay methods permits a semi-quantitative assessment of the free beta-subunit. The numerical results may not reflect the actual concentration of free beta-subunit in that two different immunoassay methods are used.     

Physiological range of human chorionic gonadotropin for support of early human pregnancy.

OBJECTIVE: To determine the physiological range of hCG in early pregnancy. DESIGN: Retrospective study of patient charts. SETTING: Magee-Women's Hospital IVF clinic, Monroeville, Pennsylvania. PATIENT(S): Sixty patients with successful, singleton birth outcomes. INTERVENTION(S): Serum hCG measurements on days 12-16 post-oocyte retrieval (OR). MAIN OUTCOME MEASURE(S): Lowest values, highest values, mean values, quartile mean values, and 48-hour doubling times for days 12-16 post-OR. RESULT(S): The average production of hCG in successful pregnancies is roughly 4-fold greater than the lowest amount observed in successful pregnancies, suggesting that a considerable excess of hCG is normally produced. Additionally, the average doubling time is almost 2-fold greater than the slowest doubling rate. CONCLUSION(S): The data from this study provide a set of values for the minimum and maximum threshold of hCG for days 12-16 post-OR that may be physiologically required, although not entirely predictive, for a successful IVF pregnancy outcome.     

A simple and sensitive nonradioactive method for the detection of urinary human chorionic gonadotropin and diagnosis of early human pregnancy. II. Single-unit test.

A simple, sensitive, and reliable single-unit nonradioactive method for the detection of human chorionic gonadotropin (hCG) in concentrated urine and the diagnosis of early pregnancy is reported. This unit, presently termed the Ayerst pregnancy test kit (APTK), consists of four components: a sampler-filter paper cone, an ultrafilter-concentrator to which a vial holder is attached, a support stand with a mirror, and an immunologic reagent vial. In the APTK, 5 to 6 ml of urine were sampled, filtered, and concentrated, and the hCG in the retentate was detected by Ayerst immunologic reagents [APTK(AY)] and by the Pregnosticon "All In" [APTK(P)]. Some of the unconcentrated urine samples (0.1 ml) were also tested in hemagglutination inhibition tests (HIT) using Ayerst [HIG(AY)] and Pregnosticon "All In" [HIT(P)] reagents. Urine samples from pregnant, nonpregnant (ovulating and nonovulating), perimenopausal, and menopausal women were tested. It was found that the APTK(AY) and APTK(P) were significantly more sensitive and reliable than the HIT(AY) and HIT(P) in detecting low levels of urinary hCG for early diagnosis of pregnancy. The sensitivity and specificity of the APTK(AY) were better than those of the APTK(P). The APTK(AY) give significantly more correct positive and negative results than the other tests performed simultaneously. The APTK(AY) is simpler and safer than the serum radioimmunoassays and radioreceptor assay presently used to detect low levels of hCG for the early diagnosis of pregnancy and other hCG-producing states.     

A simple and sensitive nonradioactive method for the detection of urinary human chorionic gonadotropin and diagnosis of early human pregnancy. I. Multiple-unit test.

A simple, sensitive, and reproducible method for the detection of urinary human chorionic gonadotropin (hCG) and diagnosis of early human pregnancy is reported. A 5-ml aliquot of filtered early-morning urine sample was concentrated in a microconcentrator (M) to 0.1 ml of retentate which was diluted with 0.4 ml of distilled water and tested in a hemagglutination inhibition test (M-HIT). Also, a 0.1-ml aliquot of filtered unconcentrated urine sample was diluted with 0.4 ml of distilled water and tested in the same hemagglutination inhibition test (HIT). Urine samples from women of reproductive age; from perimenopausal, menopausal, and proteinuric women; and from adult males were tested in the HIT and M-HIT. Some of these urine samples were also tested in the mouse ovulation bioassay (MOB). The M-HIT was significantly more reliable than the HIT for diagnosis of early pregnancy 25 to 55 days after menses. Correct negative results with the M-HIT were obtained in urine samples of most of the nonpregnant cycling, perimenopausal, and menopausal women, and adult males. Urine samples from subjects with severe proteinuria gave false-positive types of reactions in the M-HIT. Positive results were obtained in the MOB with a number of urine samples from pregnant, perimenopausal, and menopausal women. A properly conducted M-HIT should be very valuable in diagnosing pregnancy as early as the 26th day of the cycle in regularly menstruating women.     

Prenatal screening for Down syndrome with maternal serum human chorionic gonadotropin levels

Human chorionic gonadotropin levels in midtrimester pregnancies may be predictive of Down syndrome. A commercially available enzyme immunoassay kit was used to measure the beta-subunit of human chorionic gonadotropin in maternal sera from 38 Down syndrome pregnancies and 114 gestational age matched controls. The human chorionic gonadotropin levels were also assayed in 236 normal sera and plasma samples to determine normative values and appropriate individual corrections. Serum and plasma human chorionic gonadotropin levels are closely correlated and are stable at room temperature, during refrigeration, and throughout freeze-thaw cycles. There is no correlation between the human chorionic gonadotropin level and maternal age, weight, or race. However, the human chorionic gonadotropin level decreases with each week of gestation from 15 to 19 weeks. Medians for each week of gestation were established to account for this variable. Up to 63% of the Down syndrome pregnancies were detected with a cutoff of 2.0 multiples of the normal median. A computational combination of human chorionic gonadotropin and maternal serum alpha-fetoprotein testing will detect additional Down syndrome pregnancies and decrease the false-positive rate. The measurement of human chorionic gonadotropin appears to be a valuable addition to maternal serum alpha-fetoprotein screening programs that can significantly increase the proportion of Down syndrome cases diagnosed.     

Maternal serum screening for down syndrome by using alpha-fetoprotein and human chorionic gonadotropin in an asian population. a prospective study

The purpose of the present study is to evaluate the efficacy of second-trimester maternal serum screening program by using alpha-fetoprotein (AFP) and total human chorionic gonadotropin (hCG) in an Asian population. During June 1994 to July 1998, we conducted a prospective study of serum screening protocol for Down syndrome. The cut-off point for a positive result in this analysis was a risk of >/=1/270. A total of 17,742 pregnant women with singleton pregnancy were screened, and 1,153 (6.5%) had positive result. Sixteen of the 17,742 pregnancies had Down syndrome, and 10 of them had positive result. The positive rate and detective rate for Down syndrome were 6.5 and 62.5%, respectively. However, the detective rate will reduce to 47.6% after being adjusted by age-specific risk. It is indicated that the double-marker test using AFP and total hCG is an effective screen strategy for second-trimester detection of Down syndrome in Asian women.     

First-trimester maternal serum alpha-fetoprotein and human chorionic gonadotropin screening for chromosome defects.

The aim of this study was to determine the efficacy of combined maternal serum alpha-fetoprotein (MSAFP) and maternal serum human chorionic gonadotropin (MShCG) screening in detecting chromosome defects in the first trimester of pregnancy. Sera of 492 women (previously assayed for MSAFP) were analysed for MShCG under code without knowledge of cytogenetic results. Overall, 48 of 492 patients (9.8 per cent) had either an MSAFP multiple of the median less than or equal to 0.5 or an MShCG beta/alpha ratio multiple of the median less than or equal to 0.25, eight of whom had a fetus with a serious chromosome defect. A third of fetuses with Down's syndrome and 83 per cent with trisomy 18 were detected at a potential 'cost' of providing chorionic villus sampling or amniocentesis in 8.6 per cent of women screened.