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Fetal cells in maternal blood are a noninvasive source of fetal genetic material for prenatal diagnosis. We determined the number of fetal-cell DNA equivalents present in maternal whole-blood samples to deduce whether this number is affected by fetal karyotype. Peripheral blood samples were obtained from 199 women carrying chromosomally normal fetuses and from 31 women with male aneuploid fetuses. Male fetal-cell DNA-equivalent quantitation was determined by PCR amplification of a Y chromosome-specific sequence and was compared with PCR product amplified from known concentrations of male DNA run simultaneously. The mean number of male fetal-cell DNA equivalents detected in 16-ml blood samples from 90 women bearing a 46,XY fetus was 19 (range 0-91). The mean number of male fetal-cell DNA equivalents detected in 109 women bearing a 46,XX fetus was 2 (range 0-24). The mean number of male fetal-cell DNA equivalents detected when the fetus was male compared with when the fetus was female was highly significant (P = .0001). More fetal cells were detected in maternal blood when the fetus was aneuploid. The mean number of male fetal-cell DNA equivalents detected when the fetal karyotype was 47,XY,+21 was 110 (range 0.1-650), which was significantly higher than the number of male fetal-cell DNA equivalents detected in 46,XY fetuses (P = .0001). Feto-maternal transfusion of nucleated cells appears to be influenced by fetal karyotype. The sixfold elevation of fetal cells observed in maternal blood when the fetus had trisomy 21 indicates that noninvasive cytogenetic diagnosis of trisomy 21 should be feasible.

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We report a case of third trimester spontaneous resolution of a nuchal cystic hygroma with pleural effusion in a fetus with Turner syndrome (45,X). The neonatal health of the infant was good. Our case description exemplifies difficult counseling issues involved with the detection of a cystic hygroma and provides documentation of resolution late in pregnancy.

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OBJECTIVE: The purpose of this prospective study was to assess the value of maternal serum screening between 11 and 15 weeks of gestation to detect fetal Down syndrome. STUDY DESIGN: Blood samples were collected on 993 women between 11 and 15 weeks' gestation before amniocentesis. Ninety percent were > or = 35 years old. Samples were coded and assayed for alpha-fetoprotein, unconjugated estriol, and human chorionic gonadotropin. Medians were established at each week between 11 and 15 from 836 normal, singleton pregnancies. RESULTS: We used a computer-generated cut-off risk for Down syndrome of one in 365 at term; nine of 11 (82%) Down syndrome pregnancies were identified. There were 23% false-positive results in women > or = 35 years old and 6% in those < 35 years. CONCLUSION: These results suggest that maternal serum screening between 11 and 15 weeks may provide an acceptable alternative to screening between 16 and 20 weeks.

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OBJECTIVE: It is the purpose of this report to evaluate our experience with amniocentesis at less than or equal to 12 weeks' gestation. STUDY DESIGN: Medical records of 936 patients at less than or equal to 12.8 weeks' gestation undergoing genetic amniocentesis between Oct. 1, 1986, and June 30, 1990, were evaluated for gestational age, indication, frequency of needle insertion, amniocentesis complications, and pregnancy outcome. RESULTS: There were seven miscarriages within 2 weeks of amniocentesis (0.7%), 21 miscarriages before 28 weeks (2.2%), and four stillbirths or neonatal deaths (0.4%), resulting in a total postprocedural loss rate of 3.4%. There were 26 chromosomally abnormal fetuses (2.8%). The spontaneous abortion rate in ultrasonographically normal pregnancies at less than 14 weeks, not undergoing amniocentesis, has been estimated at 2.1% to 3.2%. CONCLUSION: Amniocentesis at 12 weeks is a viable option for patients desiring earlier prenatal genetic diagnostic information.

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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.

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A total of 681 pregnant women were referred for evaluation of elevated maternal serum alpha-fetoprotein levels. Ultrasonographic examination yielded an explanation for the elevation of maternal serum alpha-fetoprotein in 42% of patients. Diagnoses made by ultrasonography included incorrect fetal dating, multiple gestation, fetal death, open neural tube defect, abdominal wall defect, placental abnormalities, cystic hygroma, renal anomalies, and oligohydramnios. Optimal prenatal diagnosis of fetal anomalies also requires the use of amniocentesis in many patients. Amniocentesis may be obviated if fetal dating is incorrect, if an unsuspected multiple gestation is discovered, or if there is a clear anomaly and the parents do not desire genetic counseling based on karyotype information. If the fetus appears normal, the ultrasonographic results are equivocal, or the parents desire more detailed genetic counseling when an anomaly is found by ultrasonography, then amniocentesis should be performed. Thirteen abnormalities were diagnosed by amniocentesis alone in this group.

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Between October 1, 1986, and September 30, 1987, 1721 amniocenteses were performed at the University of California, Davis, Medical Center. Of these procedures 527 (30.6%) were early amniocenteses. Medical records were reviewed for maternal age, amniocentesis indication, color of amniotic fluid, gestational age, frequency of needle insertion, complications of amniocentesis and delivery, results of prenatal testing, and pregnancy outcome. Complete follow-up data were available for 517 (98.1%). There were 10 miscarriages before 28 weeks' gestation (1.9%), one loss after 28 weeks (0.2%), and one stillbirth (0.2%), resulting in a total postprocedural loss rate of 2.3%. Miscarriage within 2 weeks of amniocentesis occurred in four subjects (0.8%).

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Cervical mucus was collected from 35 women after artificial insemination. Mucus collections were performed at 1 h, 1 day, 2 days, or 3 days following insemination. Sperm viability was greater than 80% at all recovery times as assessed by exclusion of the supravital dye Hoechst 33258. Virtually 100% of the viable sperm were acrosome-intact at all times as assessed with a fluorescein isothiocyanate-conjugated pea lectin. Sperm were recovered from the mucus after migration into the Biggers, Whittin, and Whittingham medium in vitro. Sperm did not undergo the acrosome reaction in response to human follicular fluid immediately after migration from the mucus but did respond to this agonist after 6 h of incubation in vitro. Sperm recovered at all times after insemination had the same pattern of response to follicular fluid. Sperm that penetrated a column of cervical mucus in vitro also responded to follicular fluid with an increase in acrosome reactions after migration from the mucus and incubation for 6 h in vitro. Unlike the sperm that migrated from cervical mucus, sperm that were separated from semen by Percoll density centrifugation did not undergo the acrosome reaction when challenged with follicular fluid after 6 h but did respond after 24 h incubation. Sperm that migrated from cervical mucus had a similar increase in acrosome reactions after 6 h incubation, regardless of whether the acrosome reaction agonist was follicular fluid or disaggregated human zona pellucida.(ABSTRACT TRUNCATED AT 250 WORDS)

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Calcium oxalate is the predominant constituent of most kidney stones. The rare genetic disorder, primary hyperoxaluria, is characterized by the continuous excessive synthesis and urinary excretion of oxalic acid, leading to stone formation and renal insufficiency. The earliest measurement of oxalate in suspected cases of primary hyperoxaluria is advantageous and would lend support for continued analysis and eventual confirmation of the disease. Therefore, we quantitated oxalate levels in amniotic fluid (AF) using medium pressure ion-exchange chromatography. The mean concentration of oxalate in amniotic fluid was 1.67 mg/l +/- 0.8 (SD); (range 0.64 to 5.11 mg/l). The mean oxalate/creatinine ratio (O/C) was 0.23 +/- 0.11 (SD); (range 0.07 to 0.53). This ratio is similar to that found in the urine of infants less than 1 year (0.19 +/- 0.10; n = 17). There was no significant difference between males and females in oxalate concentration or O/C ratio. Regression analysis showed no significant correlation of fetal age with oxalate, O/C or creatinine. Studies in 13 sets of di-amnionic twins showed no statistical difference in oxalate or O/C between twin A and B. This study demonstrates the ability to accurately quantitate oxalate in amniotic fluid by ion-chromatography, and suggests that this may have a potential application in the initial screening process for the prenatal detection of primary hyperoxaluria.

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The University of California Davis Medical Center has offered secondary and tertiary care to pregnant patients with abnormal maternal serum alpha-fetoprotein levels since the start of the California alpha-Fetoprotein Screening Program in April 1986. In our first year of involvement with this method of prenatal screening, 452 patients were referred to our center for further evaluation and follow-up of either high or low maternal serum alpha-fetoprotein levels, as determined by the standard multiple of the median adjusted for maternal weight, race, and presence of insulin-dependent diabetes mellitus. This article presents the first-year results of our clinical experience. Information is presented concerning medical complications, screening and diagnostic test results and their confirmation, clinical procedures performed, and postpartum follow-up reports. Findings of pregnancy outcomes associated with low and high alpha-fetoprotein levels are presented and discussed.

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Mammalian sperm must be acrosome reacted before penetrating the zona pellucida. In some species the sperm undergo the acrosome reaction before binding to the zona pellucida and in other species only acrosome intact sperm can initiate binding to the zona. In this study we addressed the question of acrosomal status and sperm-zona binding with human gametes. Sperm acrosome reactions were induced by treatment with human follicular fluid or N-(6-amino-hexyl)-5-chloro-naphthalene sulfonamide (W-7). The sperm suspensions, containing various percentages of acrosome-reacted sperm, were then incubated with human oocytes for 1 min. The acrosomal status of the sperm population bound to the zona was similar to the acrosomal status of the population of sperm in suspension (R2 = 0.77), regardless of the treatment to induce acrosome reactions. Our interpretation of these results is that both acrosome intact and acrosome-reacted human sperm can initiate binding to the zona pellucida. However, we reported earlier (N. L. Cross, P. Morales, J. W. Overstreet, and F. W. Hanson, 1988, Biol. Reprod. 38, 235-244) that the human zona pellucida is able to induce acrosome reactions. Thus, to exclude the possibility that sperm had undergone the acrosome reaction on the zona within 1 min of binding, sperm were suspended in a nominally calcium-free Tyrode's medium (0 Ca-mTyr) before incubation with oocytes (this medium was supplemented with SrCl2 and spermine to support sperm motility and zona binding). In 0 Ca-mTyr, the proportion of acrosome-reacted sperm on the zona was still highly correlated with the proportion of reacted sperm in suspension, indicating that the sperm were reacted before binding. Evidence that 0 Ca-mTyr effectively inhibited acrosome reactions induced by the zona pellucida was derived from experiments in which sperm were treated with human follicular fluid or control medium and the suspensions were diluted with either 0 Ca-mTyr or control medium.4+ Human oocytes were added for 1 min (pulse) at which time some oocytes were fixed and other oocytes were transferred to sperm-free medium and incubated for 35 min (chase) before fixation. Sperm diluted in control medium, pretreated with either human follicular fluid or control medium, showed a similar increase (40%) in the percentage of acrosome reactions among the zona-bound sperm after the chase. Sperm diluted in 0 Ca-mTyr did not show an increase in the percentage of acrosome-reacted sperm on the zona pellucida after the chase.(ABSTRACT TRUNCATED AT 400 WORDS)

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We have shown that alpha-fetoprotein levels in amniotic fluid peak at 13 weeks and then fall by about 10% a week until 20 weeks. Use of cutoff levels obtained by extrapolation backward from medians at later dates gives results that are too high and could cause diagnostic errors. The ability to monitor alpha-fetoprotein levels accurately provides an advantage to this method of early prenatal diagnosis.

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Cervical mucus is a glycoprotein gel whose biological functions depend upon its macromolecular architecture. Using freeze-substitution fixation techniques, we have used transmission electron microscopy to examine the fine structural aspects of mucus, before and after unidirectional physical shearing, and during its interaction with sperm. The microstructure of mucus that has not been directionally stretched consists of a homogenous pattern of interconnecting electron-dense elements. The thickness of the primary structural elements varies from 0.04 to 0.5 microns, giving the impression that the elements have fibrillar but also ribbon-like properties. This dimension is smaller than the sizes depicted by prior studies using scanning electron microscopy. Within the mucus interior, the interstitial distance between adjacent primary elements ranges from 0.5 to 0.8 microns, and is filled with a fibrous network of secondary structural elements. This interstitial dimension is also significantly smaller than that suggested by prior electron microscopic work. At the exterior borders of the mucus, the interstitial sizes are reduced. After physical stretching by forceps, the mucus microstructure undergoes a radical deformation. In some specimens, the primary structural elements become longitudinally aligned and are less interconnected, with compaction at the exterior mucus borders. In other specimens, the primary structural elements become laterally compressed along the longitudinal axis; within the regions of compression, the intrapore diameter is reduced to less than 0.1 microns. These compressed regions will often exclude spermatozoa. Individual sperm deform the local mucus microstructure. The mucus directly anterior to the sperm head is stretched, and the mucus adjacent to the bending flagellum is compressed on the forward side and stretched on the opposite side.

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We have used two approaches to test the ability of the human zona pellucida to induce acrosome reactions in human sperm. First, nonviable human oocytes were incubated for 1 min in a suspension of capacitated sperm (of which fewer than 5% were acrosome-reacted) to allow binding of about 200 sperm per oocyte. Some of the oocytes were fixed immediately, and the remainder were fixed after a further 1-h incubation without free-swimming sperm. As determined by light microscopy, sperm on the zona were only 3 +/- 2% (avg. +/- SD) acrosome-reacted at 1 min, and the incidence increased to 46 +/- 15% during the next hour. Electron microscopy confirmed that most sperm on the zona at 1 min were acrosome-intact. A few sperm were in an early stage of the acrosome reaction. Acrosome reactions occurring on the zona during the subsequent hour appeared to be morphologically normal. Second, treatment of sperm in suspension with acid-disaggregated zonae (2 to 4 zonae/microliter) increased the incidence of acrosome-reacted sperm from 3 +/- 1% to 24 +/- 4%. We conclude that the human zona pellucida, or material intimately associated with it, can induce acrosome reactions in human sperm.

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Congenital cystic adenomatoid malformation Type III with fetal ascites was diagnosed in a 19-week fetus after finding an abnormally elevated maternal serum alpha-fetoprotein value during routine screening. This discovery led to early elective termination of pregnancy. Sonographic evaluation of the fetal thorax is recommended in all cases referred for ultrasound because of unexplained elevation of maternal serum alpha-fetoprotein.

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A novel Y/13 translocation was discovered in a female fetus during amniocentesis. The familial translocation is present in a phenotypically normal female sibling and in the father who also possesses a pericentric inversion. Karyotype of the father is 46,XY,-13,+der(13)t(Y;13)(q12;p12),inv(22)(p13q12.1).

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The authors have developed an extension of the sperm penetration assay for detecting serum immunoglobulins to sperm antigens that are transferred to the plasma membrane of a sperm-penetrated hamster oocyte. After the hamster oocytes have been scored for sperm penetration by observing for the presence of swollen sperm heads, they are incubated in serum followed by either a 20-minute treatment with rhodamine-conjugated protein A (which binds to most subclasses of IgA, IgG, and IgM) or a 2-hour incubation in guinea pig serum (complement). Positive fluorescence indicates that the serum contains antibodies to sperm antigens that were transferred to the surface of an oocyte during gamete fusion. Complement-mediated lysis indicates that the immunoglobulin that is bound can also fix complement. The advantages of this assay for detection of serum antisperm antibodies are that it is an extension of a widely used assay, is rapid and requires readily available reagents and equipment, can detect most subclasses of IgA, IgG, and IgM, detects antibodies to those sperm antigens that may be transferred to the oocyte during fertilization, and indicates whether the detected antisperm antibodies can mediate complement-dependent lysis of the fertilized oocyte.

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