Benachi 2015.

Study characteristics
Patient sampling	Study design: blinded, retrospective analysis from a prospective cohort. Participants: all pregnant women considered at high risk of fetal aneuploidies who were willing to undergo invasive procedure. Inclusion criteria: at least 18 years old, more than 10 weeks of gestation and singleton or twin pregnancies. Exclusion criteria: vanishing twin or < 18 years old.
Patient characteristics and setting	Number enrolled: 900 pregnant women. Number available for 2 x 2 table: 886 pregnant women (subgroup of 98%). Setting: 29 centres. French Fetal Medicine Centres in France. Recruitment period: December 2012 to October 2013. Ethnicity: Caucasian (84.2%), Black or Caribbean (4.6%), Asian (2.0%), mixed (5.7%) and unknown (3.5%). Median gestational age (range): 15.1 (10.2 to 34.6) weeks. Median maternal age (range): 35 (30 to 39) years. Relevant tests carried out prior to index test: ultrasonography (nuchal translucency measurement) and biochemical screening. Language of the study: English.
Index tests	gNIPT by MPSS with Illumina v3 flow‐cell on a HiSeq 1500 sequencer in 12‐plex. Mean fetal fraction DNA: group 1 (patients without abnormal fetal ultrasound findings, but at high risk of fetal aneuploidy): 10.9% and group 2 (high risk of fetal aneuploidy after ultrasound finding): 11.2%. Blood samples for gNIPT were collected just before reference standard. Cutpoint: positive if Z score > 3 (T21) or > 3.95 (T18 and T13). Commercial test: Laboratoire CERBA's prenatal test.
Target condition and reference standard(s)	Target conditions: T21, T18 and T13. Reference standards: fetal karyotype of chorionic villi or amniotic fluid or neonatal clinical examination.
Flow and timing	Blood samples were obtained prior to the invasive procedure (reference standard). gNIPT was a second‐tier test. 8/900 samples without karyotype result were excluded. 42 samples failed the initial MPSS testing for technical issues. 42/42 repeated tests using a second aliquot and 36/42 samples obtained gNIPT results. 6/892 samples failed during gNIPT process (low fetal fraction DNA or result appeared atypical) (no gNIPT result).
Comparative
Aim to study	To evaluate the performance of the gNIPT (using fetal ccfDNA) for detection of the 3 main autosomal fetal trisomies in a very high‐risk population of patients whose fetuses display ultrasonographically identified anomalies by comparing the results with those obtained by conventional fetal karyotyping.
Funding source or sponsor of the study	Funding source not reported. 1 author is an employee of Laboratoire CERBA and also a shareholder.
Informations about the authors contacted	Authors were contacted on: 25 May 2016. Reply received on: 26 May 2016.
Notes	Authors are from de Collaborative SEquençage a Haut Debit et Aneuploidies (SEHDA) Study Group.
Methodological quality
Item	Authors' judgement	Risk of bias	Applicability concerns
DOMAIN 1: Patient Selection
Was a consecutive or random sample of patients enrolled?	Yes
Was a case‐control design avoided?	Yes
Did the study avoid inappropriate exclusions?	No
		High	Low
DOMAIN 2: Index Test MPSS
Were the index test results interpreted without knowledge of the results of the reference standard?	Yes
If a threshold was used, was it pre‐specified?	Yes
		Low	Low
DOMAIN 3: Reference Standard
Is the reference standards likely to correctly classify the target condition?	Yes
Were the reference standard results interpreted without knowledge of the results of the index tests?	Yes
		Low	Low
DOMAIN 4: Flow and Timing
Was there an appropriate interval between index test and reference standard?	Yes
Did all analysed patients receive the reference standard?	Yes
Were all patients included in the analysis?	No
		High