BOCHUM (Germany) – Noninvasive prenatal testing has traditionally been mostly performed by centralized testing laboratories, but several providers – including Multiplicom of Belgium and UK-based Premaitha Health – have launched CE-marked NIPT tests that allow laboratories to implement the assay locally.
Multiplicom, which was recently acquired by Agilent Technologies, launched the Clarigo test in 2015. The test assesses fetal risk of trisomy 21, 18, and 13 and determines fetal sex as an option. It uses a targeted next-gen sequencing approach that relies on the company's proprietary multiplex PCR technology, called MASTR (multiplex amplification-specific targets for resequencing). Multiplicom validated the test with three laboratories on more than 1,900 samples, initially using Illumina's MiSeq and HiSeq.
According to a company spokesperson, the Clarigo test has since been adopted for routine use by 20 laboratories in 18 European countries, and more laboratories are currently implementing the test. In a local laboratory setting, the test has been validated on the MiSeq, NextSeq, and HiSeq platforms.
At the German Society for Human Genetics (GfH) annual meeting in Bochum today, Peter Nürnberg, CEO of genomic services provider Atlas Biolabs, talked about his company's adoption of the Clarigo test and its experience so far. For this process, Berlin-based Atlas collaborated with Praxis für Humangenetik in Berlin, which offers diagnostic genetic testing services to patients.
Atlas, the first laboratory in Germany to implement the Clarigo test, runs the assay on an Illumina MiSeq, which allows multiplexing of up to 12 samples. Nürnberg said being able to run the assay on a low-throughput sequencer lowers the threshold for even small laboratories to adopt it. Another advantage of the test, he said, is that Multiplicom offers it in conjunction with a dedicated data analysis tool, Clarigo Reporter, so labs do not need extensive in-house bioinformatics expertise. The sequencing data is uploaded to Multiplicom's server for the analysis.
At Atlas, the test currently takes a little more than two days to perform, including DNA extraction, amplification, library preparation, sequencing, and data analysis, although it would be possible to streamline this further, he said, as the test alone only takes about eight hours.
The implementation process started with visit from Multiplicom, followed by hands-on training of Atlas' staff at Multiplicom. After that, Atlas ran 12 proficiency samples that were provided by Multiplicom and consisted of genomic DNA rather than DNA from pregnancy samples.
On its first "real" run, three samples had "non-automated calls" or NACs, meaning the software could not call some of the results. The reason for this was that these samples did not have the required minimum depth of 2 million sequence reads, Nürnberg said, a technical problem that did not occur in later runs.
In the next run, all samples had sufficient depth of coverage, but one sample had a fetal DNA fraction of less than 4 percent, which is the cutoff for the Clarigo Reporter software, so no results were reported for that sample.
The assay does require some operator experience: Nürnberg said that when Atlas had a new technician perform the test for the first time, three samples came back with a "low fetal fraction" result, which in this case was due to inconsistent sample preparation by the operator.
The assay is also sensitive to sample quality, he said. In one case, Atlas processed a blood sample that was several days old, which resulted in test failure because of low fetal fraction.
Atlas compared results from 36 samples, 18 of which were from women with increased fetal trisomy risk, to results from Ariosa's Harmony test, which Atlas had performed on the same samples in parallel, and found no discordant results except for the one failed test from the old blood sample. All tests were conducted on samples taken between 11 and 20 weeks of pregnancy.
Nürnberg also reported results from another testing laboratory, Cytogenetics Lab Brno in the Czech Republic, which has been using the Clarigo test for one year, performing it on 343 samples so far. Using NIPT has resulted in fewer invasive diagnostic tests at that lab – 1,000 per year instead of 1,200 per year before. Overall, the Brno lab has had six samples that were positive for trisomy 21, one that was positive for trisomy 13, and two positives for trisomy 18. One of the T18 results was a false positive, though, resulting from a technical error that was corrected on repeat testing.
The Brno lab has also improved its test performance over time: initially, about 17 percent of its samples had a low fetal fraction initially but after a year, that proportion had come down to about 7 percent. Nürnberg said this has to do with how plasma is separated from the blood samples, which needs to be done correctly.
Overall, he said, it is important to follow Multiplicom's protocols for the Clarigo test to the dot and try to be consistent by using the same sequencing instrument and operating personnel.
In a follow-up conversation, Nürnberg, who is also the executive leader of the Cologne Center for Genomics at the University of Cologne, said that Atlas Biolabs plans to start offering the Clarigo test routinely in the near future.