Non-Invasive Prenatal Testing (NIPT) is a screening method that analyzes cell-free DNA (cfDNA) circulating in a pregnant person’s bloodstream to assess the risk of certain fetal chromosomal conditions. This blood test offers a non-invasive way to gain insights into a pregnancy’s genetic health, focusing on conditions like Trisomy 21 (Down syndrome), Trisomy 18, and Trisomy 13. When a pregnancy involves twins, the genetic analysis becomes more complex, introducing challenges and limitations to the test’s performance and interpretation. Understanding how NIPT works and how its capabilities are altered by a multiple gestation is important for accurate interpretation.
The Mechanics of Non-Invasive Prenatal Testing
NIPT functions by analyzing cfDNA, which consists of small fragments of genetic material released into the mother’s blood, originating from both her own cells and the placenta. Since placental cells share the same genetic makeup as the fetus, the DNA fragments derived from the placenta are used to screen for chromosomal abnormalities. The test works by quantifying the amount of DNA fragments from specific chromosomes; an extra copy of a chromosome, such as in Trisomy 21, will result in a slightly elevated count of those specific DNA fragments.
A concept known as “fetal fraction” (FF) is the percentage of total cfDNA in the maternal blood that is of placental origin. The test requires a minimum FF, typically around 4%, to yield a reliable result. This fraction generally increases throughout the pregnancy, averaging between 10% to 15% between 10 and 20 weeks of gestation. If the fetal fraction is too low, the test may fail to return a result, as there is insufficient fetal DNA to compare with the maternal DNA.
NIPT’s Capabilities in Identifying Multiples
While NIPT is not intended as a primary tool for detecting twins—ultrasound serves that function—the test can often provide an indirect indication that a multiple gestation is present. The most straightforward indicator is the overall amount of cfDNA, as the total fetal fraction (combined DNA from both placentas) is higher in a twin pregnancy than in a singleton pregnancy. This higher concentration of cfDNA can signal the presence of multiple fetuses contributing genetic material.
Another indicator is the detection of the Y chromosome, which determines the sex of the fetuses. If the test detects Y chromosome material, it confirms that at least one of the fetuses is genetically male. In cases of dichorionic (fraternal) twins, some NIPT methods can analyze genetic markers to help determine the zygosity of the fetuses. However, the test cannot definitively determine the exact number of fetuses beyond confirming the presence of multiples.
Accuracy and Limitations of Aneuploidy Screening in Twin Pregnancies
The accuracy of NIPT for aneuploidy screening is lower in twin pregnancies compared to singleton pregnancies, primarily due to the “pooled DNA” problem. When screening for conditions like Trisomy 21, the test analyzes a single, combined sample of cfDNA from all fetuses. This means the NIPT cannot distinguish which twin contributed the DNA fragment, making it impossible to identify which fetus is affected if an abnormality is detected.
The lower Positive Predictive Value (PPV) in twins means a high-risk result is more likely to be a false positive than in a singleton pregnancy. A confounder is the “Vanishing Twin Syndrome,” where one embryo stops developing early in the pregnancy. If that non-viable twin had a chromosomal abnormality, its residual placental DNA can persist for weeks, causing a false positive result for the surviving twin. Furthermore, the test’s reliability is complicated by chorionicity, as identical (monochorionic) twins share a placenta, which affects the distribution of cfDNA compared to fraternal (dichorionic) twins.
When NIPT Indicates Multiples: Follow-Up Procedures
A high-risk NIPT result in a twin pregnancy serves only as a screening indicator, not a definitive diagnosis. Any result suggesting an increased risk for an aneuploidy requires confirmation through further testing. The immediate next step is typically a high-resolution ultrasound, used to confirm the number of fetuses, determine their chorionicity (whether they share a placenta), and check for structural anomalies.
If the NIPT returns a high-risk result, the healthcare provider will recommend diagnostic testing to confirm the finding and identify the affected fetus. These invasive procedures include amniocentesis or chorionic villus sampling (CVS), which involve collecting cells directly from the amniotic fluid or placenta. Because these procedures carry a small risk of complication, they are reserved for cases where the screening test indicates risk. The decision to proceed with diagnostic tests is made after thorough genetic counseling.