Pergament 2014.

Study characteristics
Patient sampling	Study design: blinded, prospective cohort study. Participants: pregnant women from a population with mixed risk of fetal aneuploidy presenting for aneuploidy screening (51% high risk and 49% low risk). Inclusion criteria: women were 18 years of age or older with a singleton pregnancy of at least 7 weeks of gestation and signed an informed consent. Exclusion criteria: women with confirmed sex chromosome abnormality (47,XXX, XXY, XYY), confirmed triploidy, confirmed fetal mosaicism or multifetal pregnancy or egg donor.
Patient characteristics and setting	Number enrolled: 1064 pregnant women. Number available for 2 x 2 table: 963 pregnant women for T21, 964 for T18 and 45,X and 965 for T13 (subgroup of 91%). Setting: 35 centres. Prenatal care centres worldwide in Czech Republic, Japan, USA, Ireland and Spain. Recruitment period: not reported. Ethnicity: not reported. Mean gestational age (± SD; range): 17.0 (± 4.1; 7.6 to 40.6) weeks. Median gestational age: 14.3 weeks. Mean maternal age (± SD; range): 30.3 (± 7.4; 18 to 47) years. Median maternal age: 30.0 years. Relevant tests carried out prior to index test: ultrasonography (nuchal translucency measurement) or biochemical screening or both. Language of the study: English.
Index tests	gNIPT by TMPS (SNP‐based method) on Illumina Genome Analyzer IIx or HiSeq sequencers, 19,488‐plex targeted PCR with NATUS algorithm. Range fetal fraction DNA: 2% to 50%. Blood samples for gNIPT were collected before (93%) or 4 days or later after (7%) reference standard. Cutpoint: not reported. Commercial test: Natera's prenatal test.
Target condition and reference standard(s)	Target conditions: T21, T18, T13 and 45,X. Reference standards: fetal karyotype with confirmatory fluorescence in situ hybridisation or cytogenetic karyotype analysis or by genetic testing of cord blood, buccal sample, saliva, or products of conception, post‐natal or post‐live birth follow‐up.
Flow and timing	Blood samples for gNIPT were obtained prior (93%) or after (7%) to the invasive procedure (reference standard). gNIPT was a second‐tier test. 13/1064 samples excluded for other aneuploidies, including 6 cases with triploidy, 3 fetal mosaics, 2 cases with 47,XXY, 1 case with 47,XXX and 1 case with 47,XYY. 85/1051 samples failed quality control (no gNIPT result) including 64 low fetal fraction DNA, 12 low DNA, 6 contaminations, 2 loss of heterozygosity and 1 poor model fit. Between 1 to 3 samples did not passed quality control for all 5 chromosomes. No repeated test reported.
Comparative
Aim to study	To estimate performance of a single nucleotide polymorphism–based gNIPT (TMPS) for fetal aneuploidy in high‐risk and low‐risk populations on single venipuncture.
Funding source or sponsor of the study	Study funded by Natera, Inc. and a grant from the National Institute of Health, National Institute of Child Health and Human Development (4R44HD062114‐02). The majority of the authors are employees of Natera, Inc. and hold stock or options to hold stock in the company.
Informations about the authors contacted	Author was contacted on: 22 June 2016. No reply received from the author.
Notes
Methodological quality
Item	Authors' judgement	Risk of bias	Applicability concerns
DOMAIN 1: Patient Selection
Was a consecutive or random sample of patients enrolled?	Unclear
Was a case‐control design avoided?	Yes
Did the study avoid inappropriate exclusions?	No
		High	High
DOMAIN 2: Index Test TMPS
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