New single-cell prenatal blood test can identify genetic abnormalities

Proof of concept study assesses DNA from numerous live cells using single-cell–based droplet digital PCR assay without cell fixation, cell staining, or whole-genome amplification, reports The Journal of Molecular Diagnostics

Philadelphia, February 4, 2020

Non-invasive prenatal tests (NIPTs) are used for fetal genetic disease screening in pregnant women. In contrast, invasive tests like amniocentesis carry the risk of causing fetal harm. A report in The Journal of Molecular Diagnostics, published by Elsevier, describes the development of a single-cell DNA assessment method with high sensitivity and specificity. This noninvasive test enables direct extraction of genetic information from live fetal cells for simultaneous evaluation, thereby improving the likelihood of detecting a fetal anomaly.

Established NIPTs have important limitations including noninformative results. Therefore, novel methods for noninvasive definitive diagnosis of fetal genetic abnormalities are needed. Using a modified single-cell–based droplet digital PCR (sc-ddPCR) NIPT, researchers conducted a proof of concept study that successfully assessed the genetic information of extremely rare fetal cells in maternal peripheral blood. This modified sc-ddPCR system makes it possible to directly assess single-cell DNA information from live cells without cell-fixation, cell-staining, and whole genome amplification steps.

“Because NIPTs currently analyze small DNA fragments, it can be challenging to ascertain whether the origin of each DNA fragment is the mother or her fetus,” explained lead investigator Kenichiro Hata, MD, PhD, Chairman of Maternal-Fetal Biology at the National Research Institute for Child Health and Development, Tokyo, Japan. “We have observed that in some cases NIPT results are discordant with the fetal genetic information that has been reported. This study serves as a proof of concept for noninvasive prenatal diagnosis using circulating fetal cells without any strict cell purification.”

A) Schema of the single-cell–based droplet digital PCR (sc-ddPCR) system. Using the QX200 ddPCR system, each cell is simply encapsulated in one droplet. Up to 3,000 cells per well can be individually encapsulated. Subsequently, cell lysis and PCR with probes and primers are performed in each droplet. Finally, by quantifying the fluorescent droplets, single-cell genomic DNA is able to be assessed. B) Confirmation of single-cell encapsulation after generating droplets with a human B-cell line. Left, in bright field; right, with a Hoechst filter. C) Two-dimensional plot of signal amplitude of SRY and RPP30 probes in each droplet detected using our modified sc-ddPCR system. Signal amplitude of probes with 1× PBS as a blank control (left), a human female B-cell line (HEV0230) as a negative control (middle), and a human male B-cell line (HEV0057) as a positive control (right).

With this modified technique, up to 3,000 cells per well can be encapsulated in each droplet and simultaneously analyzed. The original sc-ddPCR system simultaneously assesses single-cell genetic information with high sensitivity and specificity. However, the original system is only useful for cell suspensions with a clear background (i.e., washed cell lines or clearly sorted cells with fluorescence-activated cell sorting). Researchers modified this sc-ddPCR system to improve the PCR environment in each droplet with higher sensitivity and specificity, which makes it possible to now assess single-cell genetic information from crudely purified nucleated cell samples with impurities.

To confirm the sensitivity of this modified sc-ddPCR system, investigators detected the genomic DNA of circulating male fetal cells in a crudely sorted cell suspension at the single-cell level derived from peripheral blood samples from mothers with male fetuses.

Investigators searched for the presence of the sex-determining region Y gene (SRY), which is responsible for the initiation of male sex determination. Analysis of 13 blood samples indicated that only circulating fetal cells from the three pregnant women carrying male fetuses tested positive for the SRY gene, unlike cells from the 10 pregnant women carrying female fetuses. This indicates that the modified sc-ddPCR system not only has high sensitivity, but also high specificity.

“In the future, by optimizing cell sorting and encapsulation, as well as generating a more effective PCR environment in each droplet, this modified sc-ddPCR system may be a breakthrough analysis method that can be applied to various research realms and possibly to clinical diagnostic testing,” commented Dr. Hata.


Notes for editors
The article is “Direct Assessment of Single-Cell DNA Using Crudely Purified Live Cells: A Proof Of Concept for Noninvasive Prenatal Definitive Diagnosis,” by Taisuke Sato, Yuki Ito, Osamu Samura, Hiroaki Aoki, Toru Uchiyama, Aikou Okamoto, and Kenichiro Hata ( ). It will appear in The Journal of Molecular Diagnostics, volume 22, issue 2 (February 2020) published by Elsevier.

Full text of this study is available to credentialed journalists upon request; contact Eileen Leahy at +1 732 238 3628 or Journalists wishing to interview the authors should contact Kenichiro Hata at

About The Journal of Molecular Diagnostics
The Journal of Molecular Diagnostics, the official publication of the Association for Molecular Pathology, co-owned by the American Society for Investigative Pathology, and published by Elsevier, seeks to publish high quality original papers on scientific advances in the translation and validation of molecular discoveries in medicine into the clinical diagnostic setting, and the description and application of technological advances in the field of molecular diagnostic medicine. The editors welcome review articles that contain: novel discoveries or clinicopathologic correlations, including studies in oncology, infectious diseases, inherited diseases, predisposition to disease, or the description of polymorphisms linked to disease states or normal variations; the application of diagnostic methodologies in clinical trials; or the development of new or improved molecular methods for diagnosis or monitoring of disease or disease predisposition.

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