The evolution of prenatal testing: how NIPT is changing the landscape in fetal aneuploidy screening

Dec. 19, 2015

The advent of non-invasive prenatal testing (NIPT) for fetal chromosomal abnormalities, or aneuploidies, has transformed the typical obstetrics practice and the prenatal care experience for many pregnant women. Also known as cell-free DNA (cfDNA) testing, NIPT has demonstrated better accuracy than conventional first-trimester screening and serum tests for the detection of fetal trisomies—aneuploidies that involve an extra chromosome—and its low false-positive rate in particular has reduced the need for more invasive, higher-risk diagnostic procedures, such as amniocentesis and chorionic villus sampling (CVS).1

As a result, NIPT screening technologies offer potential benefits in both patient care and cost management. But most studies assessing the performance of NIPT have focused on high-risk patient populations. Recent study data indicate that the benefits of NIPT extend to normal patent populations as well—and the results are particularly encouraging in the area of reducing false positives.

Fetal aneuploidy testing guidelines

The American College of Obstetricians and Gynecologists (ACOG) recommends that all women be offered aneuploidy screening during the first or second trimester of pregnancy.2 Most women are offered screening for trisomies 21 (Down syndrome), 18 (Edwards syndrome), and 13 (Patau syndrome).

The risk for aneuploidies increases with maternal age. The estimated risks of fetal trisomies 21, 18 and 13 for a 20-year-old woman at 12 weeks of gestation are approximately 1 in 1,000, 1 in 2,500, and 1 in 8,000 respectively. The risks of these aneuploidies for a 35-year-old woman at 12 weeks of gestation are approximately 1 in 250, 1 in 600, and 1 in 1,800.3

The most common prenatal screening test for fetal aneuploidy is the quad screen, or quadruple marker test, which is typically performed between 16 and 18 weeks of pregnancy, but occasionally up to week 20. Using a maternal blood draw, the quad screen examines four biochemical analytes: alpha-fetoprotein (AFP), a protein made by the fetus; human chorionic gonadotropin (hCG), a hormone made by the placenta; estriol, a hormone made by the placenta and the liver of the fetus; and inhibin A, another hormone made by the liver. In addition to screening for trisomies 21 and 18, the quad screen evaluates the likelihood of neural tube defects, such as spinal bifida and anencephaly, and abdominal wall defects.

The quad screen is easy to do, noninvasive, and inexpensive, and poses no risk of miscarriage or other pregnancy complications. The results of the quad test are evaluated along with maternal demographic information such as age, weight, gestational age, diabetic status, and race to derive a risk estimate using a mathematical model. The test correctly identifies about 80 percent of women who are carrying a baby with Down syndrome and has a false-positive rate of about five percent.4

Another common screening test, the nuchal translucency (NT) scan, uses ultrasound to measure the clear (translucent) space in the tissue at the back of the fetus’ neck. Babies with certain abnormalities tend to accumulate more fluid at the back of their neck during the first trimester, which causes this clear space to be larger than average. That measurement is typically combined with a maternal blood test to measure levels of pregnancy-associated plasma protein-A and human chorionic gonadotropin (hCG) at 11 to 14 weeks. The combined screening test has demonstrated a detection rate of 95 percent for Down syndrome.5 This combination test is more accurate than the Quad, but the NT scan is more difficult to perform correctly given the challenges inherent in accurately measuring the NT. While it is widely available as the standard of care, it is not available everywhere due to lack of access to NT-certified sonographers.

The impact of false positives

A major drawback of both the quad screen and the NT scan/combined screen is the high false-positive rate. Both of these tests use a statistical modeling algorithm that sets a five percent false-positive rate for trisomy 21. Multiplying the five percent false-positive rate by an estimated four million pregnancies per year in the United States (U.S.) yields 200,000 potential false-positive results for Down syndrome annually.

In reality, there are only about 6,000 cases of Down syndrome in the U.S. annually,6 and a busy obstetrics practice in the U.S. might only see one or two cases of Down syndrome a year. Patients with positive screening results typically undergo further, more invasive testing, and thus an expectant mother with a false-positive result may be subjected to a small risk of miscarriage and considerable anxiety before she can get confirmation that her pregnancy is, in fact, normal. Clearly, the high false-positive rate is a screening limitation that carries significant ramifications for patient care.

Closing the accuracy gap: cfDNA testing

Screening for fetal aneuploidy using cfDNA testing was introduced in 2011. Using a maternal blood draw, it can be performed as early as 10 weeks and poses no risk of miscarriage or other complications to the pregnancy. Studies using today’s commercially available options, all lab-developed tests, report sensitivity and specificity above 99 percent for the detection of Down syndrome. Some of these tests also measure and report fetal fraction—the percent of fetal DNA in the mothers’ blood—to provide an individualized risk score for each patient. Fetal fraction is the most critical quality control metric that impacts the accuracy of this testing.

However, the high-risk populations used in most studies of cfDNA testing to date are not representative of the normal pregnancy population, and thus they have not provided good statistical samples for the purpose of analyzing false-positive rates.

The NEXT study

That changed in 2015 with the publication of the results of the Noninvasive Examination of Trisomy (NEXT) study in the New England Journal of Medicine.7 In the NEXT study,8 18,955 women were enrolled, and results from 15,841 patients were available for analysis. The patients represented a general prenatal screening population (ages 18 to 48) from practices in the U.S., Canada, and Europe. This yielded a real-world demographic that is representative of the way obstetric screening is practiced rather than how it is studied in an academic setting.

The primary focus of the study was to compare the performance of cfDNA testing and standard first-trimester screening (with measurement of nuchal translucency and biochemical analytes) in risk assessment for trisomy 21. The authors concluded that “the performance of cfDNA testing was superior to that of traditional first-trimester screening for the detection of trisomy 21 in a routine prenatal population.” In the study, the cfDNA test demonstrated higher sensitivity (100 percent) and higher positive predictive value (80.9 percent) for Down syndrome than did standard screening (78.9 percent and 3.4 percent, respectively).

One of the most significant findings in the study, however, was the difference in the rate of false-positive results. The false-positive rate for Down syndrome with the cfDNA test was 0.06 percent—nearly 100 times lower than the 5.4 percent false-positive rate for standard screening. The implications of this improvement for prenatal care in the U.S. are significant.

Implementation considerations and future directions

The performance of cfDNA testing for the detection of trisomy 21 in a routine prenatal population was clearly superior to that of traditional first-trimester screening in the NEXT study. Having clear answers early in pregnancy regarding chromosomal abnormalities through the use of cfDNA testing can help reduce the frequency of invasive diagnostic procedures due to false positives and alleviate a significant amount of patient anxiety.

However, as the NEXT study authors note, careful consideration of screening methods and costs is needed before cfDNA testing is widely implemented for the general prenatal population. The benefits associated with the higher sensitivity and specificity and lower false-positive rate of cfDNA testing need to be weighed against the potential incremental cost to detect additional cases of trisomy 21 among women with negative results on standard screening.

Clinicians also need to consider overall patient expectations regarding prenatal genetic testing, such as the desire to screen for a variety of abnormalities that are not detected by cfDNA testing. They also need to explain the limitations and benefits of the available test choices to the patient in order to provide optimal prenatal care.


    1. Norwitz ER, Phaneuf LE, Levy B. Noninvasive prenatal testing: the future is now. Rev Obstet Gynecol 2013;6(2):48-62.
    2. ACOG Committee on Practice Bulletin No.77. Obstet Gynecol 2007; 109:217-27.
    3. Nicolaides KH. Screening for fetal aneuploidies at 11 to 13 weeks. Prenat Diagn 2011;31:7-15.
    4. Mayo Clinic. Test ID: QUAD quad screen (second trimester) maternal, serum. Accessed December 2, 2015.
    5. Johns Hopkins Medicine. Maternal serum screening, nuchal translucency scan and nasal bone sonogram screening for Down syndrome and trisomies 13 & 18. Accessed December 2, 2015.
    6. U.S. Centers for Disease Control and Prevention. Data and statistics: occurrence of Down syndrome. Accessed December 2, 2015.
    7. Norton ME, Jacobsson BB, Swarmy GK, et al. Cell-free DNA analysis for noninvasive examination of trisomy. N Engl J Med 2015;372:1589-1597.
    8. The NEXT study utilized the Harmony Prenatal Test, a cell-free DNA test that evaluates the risk for trisomies 21, 18, and 13. It is a lab-developed test performed by Ariosa Diagnostics, a CLIA-certified laboratory owned by Roche Diagnostics. This test has not been cleared or approved by the FDA. It is a screening, not diagnostic, test, and results should be confirmed by diagnostic testing.
Adam Wolfberg, MD, MPH, FACOG, practices obstetrics in Massachusetts and is the director of clinical effectiveness at athenahealth. Dr. Wolfberg is also a founder and chief medical officer of Mindchild Medical, a fetal EKG startup, and a founder of Bellybaloo.