Maternal serum screening for aneuploid pregnancy by alpha-fetoprotein, hCG, and unconjugated estriol.

A number of serum screening protocols are either currently in use or proposed to identify pregnant women under 35 years of age who are at increased risk for a Down syndrome fetus. It has been suggested that these same screening methods be applied to gravidas over 35 years of age to identify those women who should be offered amniocentesis. To evaluate the efficacy of screening for detection of aneuploid pregnancy in this age group, the serum samples of 34 women who underwent amniocentesis and who had confirmed fetal aneuploidy were assayed for alpha-fetoprotein (AFP), hCG, and unconjugated estriol (E3). These concentrations were compared with those of 85 women with known euploid pregnancy who underwent amniocentesis for advanced maternal age. The mean multiple of the median (MoM) for each of the three markers was significantly different from control values in cases of trisomy 21 (AFP median MoM = 0.82; unconjugated E3 median MoM = 0.77; and hCG median MoM = 1.89). These differences were not found when other aneuploidies were considered. Likelihood ratios were calculated for cases and controls and examined for their ability to predict the need for amniocentesis, based on currently recommended risk levels. There was a significant difference between the mean likelihood ratio for cases of trisomy 21 compared with that of controls (mean likelihood ratio = 13.48); there was no significant difference for other aneuploidies (mean likelihood ratio = 1.08). Of the 16 cases of trisomy 21 analyzed, four would not have been diagnosed antenatally if recommendation against amniocentesis had been made based on each woman's individual age-specific risk as modified by her likelihood ratio.(ABSTRACT TRUNCATED AT 250 WORDS)     

Maternal serum Down syndrome screening: unconjugated estriol is not useful

Study of 41 known Down syndrome cases and 441 matched controls did not confirm earlier reports that low unconjugated estriol levels can be used to detect fetal Down syndrome. Hence the obstetric community should exercise caution in using unconjugated estriol levels as a marker in prenatal Down syndrome screening.     

Down syndrome screening in women under 35 with maternal serum hCG.

Human chorionic gonadotropin and maternal serum alpha-fetoprotein (MSAFP) concentrations were measured in frozen maternal serum samples from 16 pregnancies with Down syndrome and from 614 unaffected pregnancies in women younger than 35. By using only maternal age and MSAFP level to determine Down syndrome risk, four of 16 (25%) Down syndrome pregnancies were identified, using risk estimates as a screening variable. This detection rate required performing amniocentesis on 4.8% of pregnancies with a normal outcome. By adding hCG level as a third risk parameter, ten of 16 (62.5%) Down syndrome pregnancies were detected. To achieve this detection rate, 4.7% of women under 35 would be recommended for amniocentesis. These results indicate that the estimation of risk for Down syndrome based on the addition of maternal hCG level to MSAFP level and maternal age will substantially improve the detection rate for Down syndrome, with no change in the amniocentesis rate. These findings are in agreement with other studies that suggest that hCG is a valuable addition to screening programs for Down syndrome.     

Very low alpha-fetoprotein in Down syndrome maternal serum screening

OBJECTIVE: To establish the frequency of very low maternal serum AFP and to differentiate congenital AFP deficiency from those diseases known to be associated with low AFP. METHODS: AFP values below 2 microg/L and borderline values up to 3 microg/L were retrospectively analysed in 839 773 singleton pregnancies included in a programme for routine screening of trisomy 21 maternal serum markers. RESULTS: Serum AFP was undetectable (< or =2 microg/L) in 8 cases, giving a frequency of 1/105 000. The calculated risk of Down syndrome was > or =1/250 in 5 cases. Fetal karyotype was normal. Seven of these pregnancies went to term (39-41 weeks) uneventfully, and birth weight was normal (3050-4110 g). In the 8th case, fetal death occurred at 35 weeks due to severe maternal diabetes. AFP levels between 2.1 and 3.0 microg/L were noted in 7 other cases. The calculated risk of Down syndrome was > or =1/250 in 5 cases, and fetal karyotype was normal. Pregnancies went to term in 4 cases (33-41 weeks), and birth weight was normal (3000-3380 g). In 3 cases, low hCG (<0.6 MoM) was associated with low AFP, and fetal death occurred at 15 to 16 weeks. CONCLUSION: Once technical errors have been excluded (repeat assay in a second run, calcium assayed to exclude the interference of EDTA for fluorimetric methods, dilution to exclude interfering antibodies, running on an alternative analyser, checking a second sample), very low second-trimester maternal serum AFP should prompt ultrasound examination in order to check fetal viability. Congenital AFP deficiency, an extremely rare disorder (1/100 000), should be suspected. It has no consequences for fetal and infant development, and parents should be reassured.     

Elevated maternal serum alpha-fetoprotein with low unconjugated estriol and the risk for lethal perinatal outcome

OBJECTIVE: To determine whether a combination of elevated maternal serum alpha-fetoprotein (MSAFP) and low unconjugated estriol (E3) concentration identifies pregnancies at particularly high risk for fetal abnormality or poor outcome. METHODS: Pregnancy outcomes were reviewed for women with elevated MSAFP (> or =2.0 MoM) from our database of 50,315 women who had received triple marker testing from 1993-1998. Outcomes for those with low E3 (< or =0.7 MoM) were compared with those with normal E3 (>0.7 MoM). The incidences of fetal death, neural tube defects, chromosome abnormalities, congenital abnormalities, preterm birth, small-for-gestational age (SGA), twins, and inaccurate dates were compared in the two groups using Fisher's exact test with P < 0.05 considered significant. RESULTS: Of the 50,315 women screened, 1,435 (2.85%) had an elevated MSAFP. Pregnancy outcomes were obtained in 94% of those with elevated MSAFP and 70% of all patients screened. Neural tube defects were present in 57 fetuses/infants (21 anencephalic, 29 spina bifida, 7 encephalocele) of which 46 (81%) had an elevated MSAFP. Of the 1,435 women with an elevated MSAFP, 199 (14%) had a low E3. Compared to those women with elevated MSAFP but normal E3, women with elevated MSAFP and low E3 were at significantly increased risk for fetal death (20.6% vs. 2.8%, relative risk (RR) 8.9), anencephaly (9.0% vs. 0.1%, RR 122.8) and chromosome abnormality (2.5% vs. 0.6%, RR 4.0). CONCLUSIONS: Pregnancies complicated by elevated second trimester MSAFP and low E3 are at a particularly high risk (32%) for lethal perinatal outcomes. Twins, while a common cause of elevated MSAFP, are rarely found when an elevated MSAFP is associated with low E3.     

Role of ultrasound for Down syndrome screening in advanced maternal age.

OBJECTIVE: To determine the sensitivity and false-positive rate of Down syndrome screening by use of maternal serum screen and the genetic sonogram in women > or =35 years of age. STUDY DESIGN: We searched our perinatal databases retrospectively from January 1992 to January 2000 for the following criteria: known Down syndrome fetus or newborn, advanced maternal age, and genetic sonogram from 14-24 weeks' gestation. The a priori maternal age or maternal serum screen risk was modified by likelihood ratios for ultrasound markers. Without markers the risk was reduced by 50%. The cut-off was 1:270. RESULTS: Age and maternal serum screen had a sensitivity of 90.5% and a false-positive rate of 27.1%. Age and ultrasound had a 95.2% sensitivity and 43.5% false-positive rate, whereas the combination of age, maternal serum screen, and ultrasound had a 97.6% sensitivity and a 22.0% false-positive rate. CONCLUSION: The combination of age, maternal serum screen, and ultrasound improves the sensitivity for Down syndrome detection in the advanced maternal age population.     

Preliminary estimate for the second-trimester maternal serum screening detection rate of the 45,X karyotype using alpha-fetoprotein, unconjugated estriol and human chorionic gonadotropin.

OBJECTIVE: To estimate the detection rate for 45,X pregnancies through second-trimester screening using maternal serum alpha-fetoprotein, human chorionic gonadotropin and unconjugated estriol. METHODS: Twenty-two cases of 45,X were ascertained through a cytogenetics database and an additional 51 cases were identified through publications. Serum analyte concentrations were reviewed for cases with fetal hydrops, cystic hygroma alone, and no evidence of edema. Using the statistical characteristics of this sample of affected pregnancies, computer simulations were carried out to determine the proportion of 45,X pregnancies that should be screen-positive for Down syndrome and trisomy 18. The extent to which additional cases of 45,X might be identified using a protocol specifically designed to detect 45,X pregnancies was also estimated. RESULTS: Approximately 54% of all 45,X pregnancies should be identifiable through screening for Down syndrome and trisomy 18. The detection rate for cases with hydrops and/or cystic hygroma was 60%, and without edema 33%. If offered with screening for Down syndrome and trisomy 18, 45,X screening could identify approximately 7% more of the affected pregnancies with an incremental rise of 0.2% in the false-positive rate. CONCLUSIONS: A screening algorithm for 45,X could be developed. However, the number of additional affected pregnancies identified would appear to be too small to justify this screening.     

Combining maternal age and serum alpha-fetoprotein to predict the risk of Down syndrome

Twenty-eight cases of Down syndrome with maternal serum alpha-fetoprotein (AFP) measured at second trimester were added to 137 previously reported cases. Seventy-eight percent of affected pregnancies had levels at or below the median AFP value, compared with 50% of unaffected pregnancies. Maternal age at delivery and serum AFP levels by multiples of the median were used to construct a table to determine the risk of Down syndrome. This was compared with the risk for a 35-year-old woman (one in 365). Similar risks were estimated for a 21-year-old with AFP below 0.4 multiples of the median, a 23-year-old below 0.5, a 26-year-old below 0.6, and a 29-year-old below 0.7 multiples of the median. This study provides guidelines for counseling pregnant women who have low serum AFP levels.     

First-trimester maternal serum unconjugated oestriol and alpha-fetoprotein in fetal Down's syndrome.

Maternal sera (MS) taken from 1396 women prior to chorionic villus sampling at 9-12 menstrual weeks were assayed for unconjugated oestriol (uE3) and alpha-fetoprotein (AFP). Median levels increased by 41 and 26 per cent per week respectively in normal pregnancies. There were 32 pregnancies with a chromosome abnormality. The median MS uE3 and AFP were 0.73 and 0.75 multiples of the median (MoM) respectively in the ten cases of Down's syndrome (DS) but not decreased in the other abnormalities. These results suggest that both uE3 and AFP may be useful in identifying DS in the first trimester. Additional prospective studies are needed to confirm these findings.     

Maternal serum alpha-fetoprotein, age, and Down syndrome risk

Before the discovery that an association existed between low maternal serum alpha-fetoprotein levels and Down syndrome, the ability to detect Down syndrome pregnancies was limited to women aged greater than or equal to 35, whose individual risks were sufficiently high (greater than or equal to 1:270 in the second trimester) to justify offering amniocentesis. If such women were to opt for that procedure, about 20% of all cases of Down syndrome could be detected. It is now possible to identify an additional 20% of all Down syndrome pregnancies in women under age 35, with the use of a screening process that combines a woman's maternal serum alpha-fetoprotein level and her age. We present a method whereby a woman's individual odds for Down syndrome can be calculated by combining the maternal serum alpha-fetoprotein measurement with her age, on the basis of published age-related Down syndrome risk data and on maternal serum alpha-fetoprotein distributions for unaffected and Down syndrome pregnancies. These individual odds calculations provide the basis both for establishing maternal serum alpha-fetoprotein/Down syndrome screening cutoffs and for counseling.     

Cost analysis of Down syndrome screening in advanced maternal age

Objectives: Fetal pulse oximetry was performed during labor in high-risk cases for fetal distress to determine the diagnostic value of this method. Methods: The fetal SpO₂ values were blinded from the obstetrician so that these values did not influence clinical decisions. Mean and lowest SpO₂ measurements for the last 30 min prior to either fetal scalp blood sampling or delivery were correlated with scalp pH or pH from the umbilical artery. Results: No significant correlation was found between pH and mean fetal oxygen saturation (correlation coefficient -0.02, p = 0.9). There was no significant correlation between pH and lowest fetal oxygen saturation (correlation coefficient 0.04, p = 0.84). Concerning the feasibility of the method, we found that only 23 of 65 included patients were suitable for analysis; in 20% of cases, we were not able to perform a SpO₂ measurement. Conclusions: None of three cases with pH below 7.05 would have been detected using mean SpO₂ over the last 30 min prior to fetal scalp blood sampling or delivery. Only one case would have been detected using the lowest SpO₂ measurement over this period. We conclude that fetal SpO₂ measurements during labor are of poor diagnostic value, with a disappointing feasibility and therefore are not ready for implementing into daily clinical practice.     

Prenatal screening for chromosome abnormalities using maternal serum chorionic gonadotrophin, alpha-fetoprotein, and age.

Human chorionic gonadotrophin (hCG) levels were assayed retrospectively in stored maternal serum samples from 78 chromosomally abnormal pregnancies and 410 controls matched for gestation and maternal age. The median serum hCG concentration in 49 pregnancies with Down's syndrome was significantly elevated, at 2.18 multiples of the normal median. Significantly reduced hCG concentrations were found in a group of four trisomy 18 pregnancies (all less than 0.4 multiples of the median). Eight cases of unbalanced chromosome rearrangements appeared to show some lowering of hCG levels, while there was no significant difference in the levels in the cases of trisomy 13, balanced translocations, and sex chromosome abnormalities. Maternal serum hCG alone is a better indicator of Down's syndrome pregnancies than maternal age or maternal serum alpha-fetoprotein (AFP), either individually or in combination, and provides a further virtually independent measure of risk. On the basis of our findings, screening for Down's syndrome using hCG and AFP results combined with maternal age risks is predicted to result in a higher detection rate (57 per cent) for a lower false-positive rate (5.0 per cent) than would be attainable by combined AFP and age screening (37 per cent detection at a 6.6 per cent false-positive rate).     

Urinary screening tests for fetal Down syndrome: II. Hyperglycosylated hCG.

Hyperglycosylated hCG is a form of hCG with more complex oligosaccharide side chains. A specific immunoassay was developed to measure hyperglycosylated hCG. Levels were measured in urine samples from 1157 women between 11 to 22 weeks of gestation, undergoing genetic analysis because of advanced maternal age. Values were normalized to urine creatinine concentration and plotted against gestational age, median values were determined and multiples of the control median (MoM) calculated. The median MoM and log standard deviation (log SD) of the 1134 control samples was 1.0 and 0.47, and of the 23 Down syndrome cases was 7.8 and 0.48, respectively. This indicated a 7.8-fold increase in hyperglycosylated hCG levels in Down syndrome cases. In the accompanying article, a stability problem was found with beta-core fragment measurements in frozen urine samples. In anticipation of similar problems, nine urine samples were tested for hyperglycosylated hCG fresh and after storage in the freezer. No clear difference was found in hyperglycosylated hCG values. In addition, no trend was found in hyperglycosylated hCG MoM values or in Down syndrome detection rates in urine samples stored for one, two or three years in the freezer. Samples were split into five equal groups according to creatinine concentration. A trend was observed, hyperglycosylated hCG MoM values decreasing with advancing creatinine concentration (1.77, 1.08, 1.01, 0.73 and 0.60 at 0.25, 0.50, 0.79, 1.11 and 1.73 mg/ml, respectively). An error was noted. This was corrected with a regression equation. After correction, the median MoM and log SD of the control samples was 1.0 and 0.44, and of Down syndrome samples was 7.3 and 0.42, respectively. Correction of this error, while reducing the elevation of Down syndrome cases, tightened the spread of samples. Samples were ranked and centiles determined. 18 of 23 Down syndrome cases (78 per cent) exceeded the 95th centile of the control population. ROC analysis indicated 79 per cent detection at 5 per cent false-positive rate. Urine samples were collected during two periods of gestation, an early period (11th to 14th completed week) and the period when chemical screening is normally performed (15th to 21st week). ROC analysis indicated 80 per cent and 78 per cent detection rates, respectively, at 5 per cent false-positive rate, in the two gestational periods. Hyperglycosylated hCG values were modelled with beta-core fragment values, total oestriol values and maternal age. ROC analysis indicated 97 per cent detection rate at 5 per cent false-positive rate. This detection rate and this level of Down syndrome and control patient discrimination surpasses that of any other serum, urine or ultrasound screening protocol. Hyperglycosylated hCG should be considered as a new screening test for aneuploid pregnancies, with the potential of detecting almost all cases of Down syndrome. Evaluation is needed by other centres in order to bring hyperglycosylated hCG into clinical practice.     

Screening for fetal Down syndrome with maternal serum hCG and oestriol: a prospective study.

We report the preliminary results of a prospective study aimed at evaluating the effectiveness of Down syndrome (DS) screening using second-trimester measurement of maternal serum human chorionic gonadotrophin (hCG) and unconjugated oestriol (uE3) together with maternal age. Reference values for hCG, uE3, and the hCG/uE3 ratio in normal pregnancies were established from more than 3000 normal gestations and found to follow a log-normal statistical distribution. Risk evaluation was made using reference values for affected pregnancies from retrospective studies. Screening of 10,000 women under 38 years resulted in 412 amniocenteses and the prenatal diagnosis of six cases of DS, whereas four cases remained undetected until term. In a parallel study, diagnostic amniocentesis was performed in women over 38 years and in women with a previous affected child, and an evaluation of the risk of fetal DS based on serum hCG and uE3 levels was made in all cases. Fourteen cases of DS were detected. Median values for hCG and uE3 in the 24 affected pregnancies were close to the 90th and tenth centiles of the normal reference values, respectively, and thus are in good agreement with the values reported by others in retrospective studies.     

First trimester biochemical screening for Down syndrome: free beta hCG versus intact hCG

To compare free beta hCG versus intact hCG in first trimester Down syndrome screening we analysed 63 cases of Down syndrome and 400 unaffected control pregnancies between 10 and 13 weeks' gestation. The Down syndrome median multiple of the median (MoM) was significantly higher (p=0.001) for free beta hCG (1.89 MoM) than for intact hCG (1.37 MoM). Although distributions for free beta hCG (unaffected, 0.2157; DS, 0.2322) are wider than for intact hCG (unaffected, 0.1697; DS, 0.2158), overall 27% of Down syndrome cases were above the 95th percentile for free beta hCG compared to 19% for intact hCG. Combined with maternal age, free beta hCG detected 45% of Down syndrome pregnancies at a 5% false positive rate. Intact hCG combined with maternal age demonstrated a detection efficiency comparable to maternal age alone (35% versus 32%). In contrast, a recent study (Haddow et al., 1998-NEJM 338: 955-961) indicated that intact hCG yielded a higher first trimester Down syndrome detection efficiency than free beta hCG (29% versus 25% respectively). Re-analysis of distribution parameters in the Haddow et al. study, however, show that free beta hCG was actually the better marker (23% detection for intact hCG versus 29% for free beta hCG).