NIPT - BeSHG

NIPT
Non-Invasive Prenatal Testing
Ben Caljon, NGS platform (UZ Brussel, VUB, ULB)
Overview
Summary/introduction on invasive testing
History of prenatal diagnosis
Current first trimester screening strategy
Types of Invasive Prenatal Diagnosis (IPD)
Assessing IPD results
Non-invasive testing
Detection of cfDNA
NIPT methodologies
NIPT technique (Lo et al)
Claimed accuracy
Indications and limitations
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NIPT - Non-invasive prenatal testing
09-05-2014
Invasive testing
Summary/introduction
History (1)
Prenatal diagnosis
1970 to early 1980 : advanced maternal age
(+35y)
<1/3 of Down syndrome pregnancies diagnosed
2% of IPD had chromosomal abnormalities
Data from England and Wales in the period of 1990-1998
Morris JK, Mutton DE, Alberman E. Revised estimates
of the maternal age specific live birth prevalence of
Down's syndrome. J Med Screen. 2002;9(1):2-6
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NIPT - Non-invasive prenatal testing
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History (2)
Late 1980, early 1990 : introduction of
second-trimester maternal serum markers
Ca. 2/3 of Down syndrome pregnancies diagnosed
4% of IPD had chromosomal abnormalities
Marker
Full name
Origin
Tr 21
AFP
alphafetoprotein
Yolk sac/fetal
liver
↓
(MoM : 0,73)
HCG
beta-human
chorionic
gonadotropin
Trophoblast
↑
(MoM : 2)
UE3
unconjugated
estriol
Feto-placental
unit
↓
(MoM : 0,73)
Marker levels vary with gestation week
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NIPT - Non-invasive prenatal testing
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History (3)
Complex data analysis
Necessary to use MoM (multiple of median) values
Need for accurate population parameters
Need for likelihood ratio distribution
Need for correct curve fitting for medians of the markers
Need for correction for other factors such as maternal weight
Software prediction tools available
MoM =
6
Patient value
Median value normal pregnancies
NIPT - Non-invasive prenatal testing
09-05-2014
History (4)
Late 1990, early 2000 : combined first-trimester
testing
Ca. 90% of Down syndrome pregnancies diagnosed
6% of IPD had chromosomal abnormalities
Marker
Full name
Origin
Tr 21
HCG
beta-human chorionic
gonadotropin
Trophoblast
↑
(MoM : 1,8)
PAPP-A
Pregnancy-associated
plasma protein A
trophoblast and decidualized endometrial
stroma cells
↓
(MoM : 0,4)
Ultrasound measurement
NT
7
Nuchal translucency
thickness
NIPT - Non-invasive prenatal testing
↑
(MoM : 2-2,5)
09-05-2014
History (5)
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NIPT - Non-invasive prenatal testing
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History (6)
Normal
Disadvantage : inter-operator differences
Trisomy 21
Advantage : useful in twins, elevated in other trisomies
Fluid filled space (cardiac failure, structural malformation,
delayed/abnormal development of lymphatic system)
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NIPT - Non-invasive prenatal testing
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History (7)
Factors influencing 1st trimester testing :
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NIPT - Non-invasive prenatal testing
09-05-2014
Current first trimester screening
Curtosy Dr. Anniek Vorsselmans :
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NIPT - Non-invasive prenatal testing
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Prenatal testing – Invasive (1)
Chorion Villus Sampling
(CVS)
10-13 weeks after gestation
Transabdominal or
Transcervical (depending on
access to placenta/skills
physician)
Risk :
Vaginal bleeding/spotting
Chorioamnionitis
Fetal loss: 1/200
Pro :
Results are available earlier
in pregnancy
Less parental anxiety
Earlier/safer methods for
pregnancy termination
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NIPT - Non-invasive prenatal testing
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Prenatal testing – Invasive (2)
Amniocentesis
>14 weeks after gestation
(earlier possible but higher risk)
Risk
Transient vaginal spotting:
1-2%
Amniotic fluid leakage: 12%
Chorio-amnionitis: < 0.1%
Needle injuries to the fetus
are rare when
ultrasonographic guidance
is used.
Fetal loss: 1/200
(overestimate)
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NIPT - Non-invasive prenatal testing
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Prenatal testing – Invasive (3)
Cordocentesis
>17 weeks after gestation
(mostly > 20 weeks)
Only if CVS or AC not possible
Risk (similar to AC)
Transient vaginal spotting:
1-2%
Amniotic fluid leakage: 12%
Chorio-amnionitis: < 0.1%
Needle injuries to the fetus
are rare when
ultrasonographic guidance
is used.
Fetal loss: 1/200
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NIPT - Non-invasive prenatal testing
09-05-2014
Assessing invasive results (1)
Preparation of samples
CVS : microscopic separation
of maternal tissue from villi =
dissection of decidua
Centrifugation of cord blood
(serum/plasma for
biochemistry, WBC for
DNA/biochemical)
Check for maternal cell
contamination (MC)
Via fragment analysis
(PowerPlex 16HS)
Dependent on analysis : put
AC/CVS cells in culture
Risk for overgrowth with
maternal cells
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NIPT - Non-invasive prenatal testing
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Assessing invasive results (2)
Example Maternal Cell Contamination
Prenatal sample
Mother
For biochemistry analysis, MC
detection is more difficult : comparison
in enzymatic activities decidua/villi
• β-glucuronidase
• β-hexosaminidase
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NIPT - Non-invasive prenatal testing
Father
09-05-2014
Assessing invasive results (3)
Biochemistry analysis
Only possible if enzyme is expressed in the
analyzed tissue (AC/CVS/cord blood)
Power : one assay for many mutations
Measure enzymatic activities via artificial
substrates (fluorimetric dosage over time)
Cytogenetics
CVS :
Direct/KT
LT
AC
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NIPT - Non-invasive prenatal testing
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Assessing invasive results (4)
Karyotype
FISH
>5 Mb dup/del
AC lower error rates than CVS :
• Confined Placental Mosaicism (CPM)
• MC
• Banding resolution
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NIPT - Non-invasive prenatal testing
QF-PCR
MLPA
aCGH (+ SNP
array)
09-05-2014
Assessing invasive results (5)
Molecular DNA analysis
Wide range of molecular test/techniques can
be applied if DNA is available
Restriction digestion
Southern blotting
Fragment analysis
Sequencing
…
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NIPT - Non-invasive prenatal testing
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Non-invasive testing
Detection of cfDNA
Detection of cell-free fetal DNA
in maternal plasma in 19971
Shedding of trophoblast cells
microparticles of fragmented DNA
maternal bloodstream short
half life (2 h clearance)
Pro : multigravid pregnancy
Median prevalence of 3% (real time
PCR) to 10%2 (digital PCR) in 1st
and 2nd trimester
Reliable detection from 11-12
weeks on
Increasing % during 2 and 3rd
trimester
1.
2.
21
Lo YMD, Corbetta N, Chamberlain PF, Rai V, Sargent IL, et al. (1997) Presence of fetal DNA in
maternal plasma and serum. Lancet 350: 485–487.
Lun FMF, Chiu RWK, Chan KCA, Leung TY, Lau TK, Lo YMD. 2008 Microfluidics digital PCR reveals a
higher than expected fraction of fetal DNA in maternal plasma. Clin. Chem. 54, 1664–1672.
NIPT - Non-invasive prenatal testing
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Introduction - Media
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NIPT - Non-invasive prenatal testing
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NIPT - Methodologies
NIPT
cfDNA based
cfRNA based
Clinical utility
s-MPS
Digital PCR
(Shotgun Massive
parallel Sequencing)
t-MPS
(Targeted Massive
parallel Sequencing)
SNP based
approaches
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NIPT - Non-invasive prenatal testing
RNA based
approaches
qPCR
• Examination of differentially
expressed genes (trophoblast
vs. Maternal RNA)
• Currently being
investigated.
• Proof-of-principle was
successful
• Clinical trial disappointing
09-05-2014
NIPT - Methodologies
NIPT
cfDNA based
cfRNA based
Clinical utility
s-MPS
Digital PCR
(Shotgun Massive
parallel Sequencing)
t-MPS
RNA based
approaches
qPCR
(Targeted Massive
parallel Sequencing)
SNP based
approaches
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NIPT - Non-invasive prenatal testing
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NIPT – Digital PCR
Absolute quantitation of nonpolymorphic chr1 (reference)
and chr21 marker.
Difference in quantitation
allows for T21 detection
Advantage :
Ease of use
Cheap
Disadvantage :
Only limited amount of markers
to be assessed in 1 experiment
Fetal fraction determines
complexity (number of
“wells”/counts) needed for
sufficient statistical power. For
2%, 220816 counts are needed,
which is beyond current
commercial technology
Lo YM, Lun FM, Chan KC, Tsui NB, Chong KC, Lau TK,
Leung TY, Zee BC, Cantor CR, Chiu RW. Digital PCR for the
molecular detection of fetal chromosomal aneuploidy. Proc
Natl Acad Sci U S A. 2007 Aug 7;104(32):13116-21.
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NIPT - Non-invasive prenatal testing
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NIPT - Methodologies
NIPT
cfDNA based
cfRNA based
Clinical utility
s-MPS
Digital PCR
(Shotgun Massive
parallel Sequencing)
t-MPS
RNA based
approaches
qPCR
(Targeted Massive
parallel Sequencing)
SNP based
approaches
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NIPT - Non-invasive prenatal testing
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NIPT – qPCR
Different array of methods,
mostly based on differentially
methylated regions (DMRs)
Methylation Dependent
Immuno-Precipitation (MeDIPPCR)
Methylation Specific PCR (MSP)
Advantage :
Cheap on consumables
Disadvantage :
Selection/optimization of test
is tedious (selection of DMRs)
Multiple DMRs needed to
obtain sufficient statistical
power
Not highly multiplexable
Laborious
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NIPT - Non-invasive prenatal testing
09-05-2014
NIPT - Methodologies
NIPT
cfDNA based
cfRNA based
Clinical utility
s-MPS
Digital PCR
(Shotgun Massive
parallel Sequencing)
t-MPS
RNA based
approaches
qPCR
(Targeted Massive
parallel Sequencing)
SNP based
approaches
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NIPT - Non-invasive prenatal testing
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NIPT – SNP based approaches (1)
Offered by company Natera
Commercial name : Panorama
NATUS technology = Next generation Aneuploidy Testing
Using SNPs
Using NGS, 11000 SNPs are determined within the cfDNA for
chromosomes 13,18,21, X and Y
PCR amplification of polymorphic regions
Sequencing of amplicons using NGS device
Based on the SNP genotyping (both plasma as buffy coat),
the NATUS algorithm performs a “per chromosome” QC and
establishes fetal chromosomal anomalies using a Bayesianbased maximum likelihood statistical method
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NIPT - Non-invasive prenatal testing
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NIPT – SNP based approaches (2)
Advantages
High accuracy (also for sex
chromosomes)
Ability to detect polyploidy
Disadvantage
Not useable for donor-egg
cell pregnancies
Only small cohort study
published (145 samples)
In proof-of-principle : 12.6%
of samples had a “no-call”
due to poor DNA quality (not
passing QC filter)
Currently in patent-conflict
with Sequenom
No information on other
chromosomes
Zimmermann B, Hill M, Gemelos G, Demko Z, Banjevic M, Baner J, Ryan A, Sigurjonsson S, Chopra N, Dodd M, Levy B,
Rabinowitz M. Noninvasive prenatal aneuploidy testing of chromosomes 13, 18, 21, X, and Y, using targeted
sequencing of polymorphic loci. Prenat Diagn. 2012 Dec;32(13):1233-41.
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NIPT - Non-invasive prenatal testing
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NIPT - Methodologies
NIPT
cfDNA based
cfRNA based
Clinical utility
s-MPS
Digital PCR
(Shotgun Massive
parallel Sequencing)
t-MPS
RNA based
approaches
qPCR
(Targeted Massive
parallel Sequencing)
SNP based
approaches
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NIPT - Non-invasive prenatal testing
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NIPT – tMPS (1)
Offered by company
Ariosa Diagnostics
Commercial name :
Harmony Prenatal Test
Scope : T13, T18, T21, X
& Y (optional)
Technology : DANSR
(Digital Analysis of
Selected Regions) assay
and FORTE (fetal-fraction
optimized risk of trisomy
evaluation)
Sparks AB, Struble CA, Wang ET, Song K, Oliphant A.
Noninvasive prenatal detection and selective analysis
of cell-free DNA obtained from maternal blood:
evaluation for trisomy 21 and trisomy 18. Am J Obstet
Gynecol. 2012 Apr;206(4):319.e1-9.
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NIPT - Non-invasive prenatal testing
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NIPT – tMPS (2)
DANSR : probes for both polymorphic
(determine fetal fraction) as non-polymorphic
regions (chromosome proportion
determination)
FORTE algoritm : calculates an individualized
odd score for trisomy, based on fetal fraction,
chromosome proportion data and maternal age
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NIPT - Non-invasive prenatal testing
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NIPT – tMPS (3)
Advantages
Easy fetal fraction determination, using probes for polymorphic regions
No need for reference data (FORTE normalizes using intra-run data) : less
process drift
Cheaper, because of higher multiplexibility
Disadvantages :
False negative results have been obtained for 1/232 T21, 2/105 T13 and 2/10
T13
No information on other chromosomes/limited normalization capabilities
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NIPT - Non-invasive prenatal testing
09-05-2014
NIPT - Methodologies
NIPT
cfDNA based
cfRNA based
Clinical utility
s-MPS
Digital PCR
(Shotgun Massive
parallel Sequencing)
t-MPS
RNA based
approaches
qPCR
(Targeted Massive
parallel Sequencing)
SNP based
approaches
35
NIPT - Non-invasive prenatal testing
09-05-2014
NIPT – sMPS (1)
Offered by companies Sequenom and
Verinata
Commercial names : MaterniT21 Plus and
Verifi resp.
Technology : see next slides
Scope
Sequenom : T21, T18, T13, X/Y aneuploidy,
T16,T22 (and “specific” microdeletions)
Verinata : T21, T18, T13, X/Y aneuploidy
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NIPT - Non-invasive prenatal testing
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NIPT – sMPS (2)
Verinata (website verinata)
N
Sensitivit 95%
y
CI
Specificity 95% CI
21
500
>99.9%
(90/90)
96100.0
99.8%
98.7-100.0
(409/410)
18
501
97.4%
(37/38)
86.299.9
99.6%
98.5-100.0
(461/463)
13
501
87.5%
(14/16)
61.798.5
>99.9%
99.2-100.0
(485/485)
MX
508
95.0%
(19/20)
75.199.9
99.0%
97.6-99.7
(483/488)
XX
XY
N
Sensitivity
95% CI Specificity 95% CI Accura 95% CI
cy
508
97.6%
(243/249)
94.899.1
99.2%
(257/259
)
97.299.9
98.4%
96.999.3
508
99.1%
(227/229)
96.999.9
98.9%
(276/279
)
96.999.8
99.0%
97.799.7
Sequenom (website verinata)
Results of published clinical
trials will be discussed later
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NIPT – sMPS (3)
Why sMPS :
False positive/negative rates decrease over time (mainly by improved
bioinformatics algorithms)
No test is perfect (tMPS, sMPS nor SNP based)
-
Numerous factors responsible for test failure / altered DRs, but mainly due to
low % fetal DNA
Natera test has not been widely validated
Applicable to egg cell donor pregnancies
Lowest result failure (retest/new sample needed)
Generic test :
Expandable to other chromosomes
Possibility to pick up subchromosomal alterations (if sequencing costs
decreases)
Possibility to determine fetal SNVs (if sequencing costs decreases)
Possibility to determine polyploidy using SNVs (if sequencing costs decreases)
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NIPT - Non-invasive prenatal testing
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Overview NIPT technique
1. Phlebotomy
5. Cluster generation
39
2. Plasma isolation
6. Sequencing
NIPT - Non-invasive prenatal testing
3. cfDNA extraction
4. Library preparation
7. Data-analysis
8. Reporting
09-05-2014
NIPT technique – Phlebotomy (1)
1. Phlebotomy
Maternal plasma + DNA isolation
Collection of maternal peripheral
blood (from 12 weeks gestation) in
10 ml EDTA tubes (Streck tubes)
with a proprietary stabilizing
agent
Inhibits gDNA release from
nucleated cells
Inhibits nuclease activity
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NIPT - Non-invasive prenatal testing
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NIPT technique – Phlebotomy (2)
gDNA release from nucleated (maternal)
cells over time (0-14 days)
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NIPT - Non-invasive prenatal testing
09-05-2014
Overview NIPT technique
1. Phlebotomy
5. Cluster generation
42
2. Plasma isolation
6. Sequencing
NIPT - Non-invasive prenatal testing
3. cfDNA extraction
4. Library preparation
7. Data-analysis
8. Reporting
09-05-2014
NIPT technique – Plasma isolation (1)
2. Plasma isolation
Centrifuge blood @ 1600g for 10’ at 4°C (Lo, 2013) /
@300g for 20’ at 22°C (Streck)
Recentrifuge plasma portion @ 16000g for 10’ at 4°C
(Lo, 2013) / @ 5000g for 10’ at 22°C (Streck)
Ca. 55% of total blood
<1% of total blood
Removal of nucleated cells, as a
source of maternal DNA
Ca. 45% of total blood
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NIPT - Non-invasive prenatal testing
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Overview NIPT technique
1. Phlebotomy
5. Cluster generation
44
2. Plasma isolation
6. Sequencing
NIPT - Non-invasive prenatal testing
3. cfDNA extraction
4. Library preparation
7. Data-analysis
8. Reporting
09-05-2014
NIPT technique – cfDNA extraction (1)
3. cfDNA extraction
Detection of cell-free fetal
DNA (cfDNA) in maternal
plasma in 19971
Shedding of trophoblast cells
microparticles of fragmented DNA
maternal bloodstream short half
life (2 h clearance)
Median prevalence of 3% (real
time PCR) to 10%2 (digital PCR)
in 1st and 2nd trimester
Increasing % during pregnancy
Reliable detection from 11-12
weeks on
1.
2.
45
Lo YMD, Corbetta N, Chamberlain PF, Rai V, Sargent IL, et al. (1997) Presence of fetal DNA in
maternal plasma and serum. Lancet 350: 485–487.
Lun FMF, Chiu RWK, Chan KCA, Leung TY, Lau TK, Lo YMD. 2008 Microfluidics digital PCR reveals a
higher than expected fraction of fetal DNA in maternal plasma. Clin. Chem. 54, 1664–1672.
NIPT - Non-invasive prenatal testing
09-05-2014
Overview NIPT technique
1. Phlebotomy
5. Cluster generation
46
2. Plasma isolation
6. Sequencing
NIPT - Non-invasive prenatal testing
3. cfDNA extraction
4. Library preparation
7. Data-analysis
8. Reporting
09-05-2014
NIPT technique – Library prep. (1)
4. Library preparation
The cfDNA has to be modified for the
sequencing instrument (eg HiSeq) to be
able to read the DNA sequences of each
individual fragment. The following actions
are required :
•
•
•
•
•
•
•
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NIPT - Non-invasive prenatal testing
End repair
Adenylation (3’)
Adapter ligation
Library purification (x2)
PCR amplification
Library purification
Library validation
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NIPT technique – Library prep. (2)
Library validation
Average size cfDNA fragments = 150170 bp
Average size cfDNA fragments +
adapters = ca. 300 bp
• Validated libraries are pooled in
equal ratios (equimolar).
• Because of the unique adapter (1
adapter per cfDNA sample), the
instrument can discriminate
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NIPT - Non-invasive prenatal testing
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Overview NIPT technique
1. Phlebotomy
5. Cluster generation
49
2. Plasma isolation
6. Sequencing
NIPT - Non-invasive prenatal testing
3. cfDNA extraction
4. Library preparation
7. Data-analysis
8. Reporting
09-05-2014
NIPT technique – Cluster gen. (1)
5. Cluster generation
The pooled library sample has to be attached
to glass slide (flowcell), so every unique
fragment binds to a random but distinct zone
on this slide
Per zone, there will be 1 unique fragment that
will be amplified, so multiple copies of the
same fragment exist on that zone (=cluster
generation)
Amplification needed to overcome current detection
limitations (not possible to detect/genotype a
single DNA fragment)
Maternal cfDNA fragment
Fetal cfDNA fragment
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NIPT technique – Cluster gen. (2)
Glass slide coated with DNA fragments
that recognize the adapters
Random binding of cfDNA fragments to probes on flowcell
Maternal cfDNA fragment
Fetal cfDNA fragment
mm
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NIPT technique – Cluster gen. (3)
Amplification of each unique
fragment per cluster (clonal
amplification)
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NIPT - Non-invasive prenatal testing
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Overview NIPT technique
1. Phlebotomy
5. Cluster generation
53
2. Plasma isolation
6. Sequencing
NIPT - Non-invasive prenatal testing
3. cfDNA extraction
4. Library preparation
7. Data-analysis
8. Reporting
09-05-2014
NIPT technique – Sequencing (1)
6. Sequencing
Each cluster is interrogated for
its sequence
Create correct single stranded template for sequencing reaction
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NIPT - Non-invasive prenatal testing
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NIPT technique – Sequencing (2)
Anneal sequencing primer
Sequencing By Synthesis (SBS)
Iteration of DNA polymerisation (1
base per cycle), laser scanning and
decapping
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NIPT - Non-invasive prenatal testing
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Overview NIPT technique
1. Phlebotomy
5. Cluster generation
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2. Plasma isolation
6. Sequencing
NIPT - Non-invasive prenatal testing
3. cfDNA extraction
4. Library preparation
7. Data-analysis
8. Reporting
09-05-2014
NIPT technique – Data analysis (1)
Resolution of detection is determined by
Coverage (no. of reads)
3 Mb : ca. 150x106 reads required
2 Mb : ca. 192x106 reads required
1 Mb : ca. 380x106 reads required
Higher resolution has high impact on total
cost
Range of currently implemented no. of reads
for detecting aneuploidies :
Min : 2x106 reads
Max : 25x106 reads
Yu et al, 2013, Noninvasive Prenatal Molecular Karyotyping from Maternal Plasma
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NIPT technique – Data analysis (1)
Raw Data
Maternal cfDNA fragment
Fetal cfDNA fragment
GC correction
Raw sequencing data is
aligned to the genome
Counting Statistics
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NIPT - Non-invasive prenatal testing
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NIPT technique – Data analysis (2)
Z-score calculation
Step 1 : normalize for amount of data
Divide no fragments for chr N with total no of
fragments
Sample 1 : 1x representation
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NIPT - Non-invasive prenatal testing
Sample 2 : 2x representation
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NIPT technique – Data analysis (3)
Z-score calculation
Step 2 : determine the number of standard
deviations from mean (control SDs should be
established for comparison purpose)
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Claimed accuracy (1)
The sensitivity/specificity of the test
varies per chromosome
Near 100% sensitivity and specificity
1.
61
Devers PL, Cronister A, Ormond KE, Facio F, Brasington CK, Flodman P. Noninvasive prenatal testing/noninvasive
prenatal diagnosis: the position of the National Society of Genetic Counselors. J Genet Couns. 2013
Jun;22(3):291-5
NIPT - Non-invasive prenatal testing
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Claimed accuracy (2)
The sensitivity/specificity of the test
varies per chromosome & with
bioinformatics corrections
1.
62
Chen EZ, Chiu RW, Sun H, Akolekar R, Chan KC, Leung TY, Jiang P, Zheng YW, Lun FM, Chan LY, Jin Y, Go AT, Lau ET, To WW, Leung WC, Tang RY, AuYeung SK, Lam H, Kung YY, Zhang X, van Vugt JM, Minekawa R, Tang MH, Wang J, Oudejans CB, Lau TK, Nicolaides KH, Lo YM. Noninvasive prenatal
diagnosis of fetal trisomy 18 and trisomy 13 by maternal plasma DNA sequencing. PLoS One. 2011;6(7):e21791.
NIPT - Non-invasive prenatal testing
09-05-2014
Claimed positive predictive value
positive predictive value, or precision
rate is the proportion of positive test
results that are true positives (such as
correct diagnoses).
reflects the probability that a positive
test reflects the underlying condition
being tested for. Its value does depend
on the prevalence of the outcome of
interest
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NIPT - Non-invasive prenatal testing
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Claimed positive predictive value
If positive predictive value, 99%
sensitivity and 0.2% false positives
If 1/50 risk
90.8%
If 1/500 risk
49.7%
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Indications
“NIPT shall replace amniocentesis”
Guidelines of HC
See next slide for indications
RIZIV
No RIZIV refund for NIPT
Biology
NIPT will fail in certain situations : twins,
translocations, subchromosomal abberations,
placental mosaicism,…
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NIPT - Non-invasive prenatal testing
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Indications
Previous indications for invasive
NIPT
Abnormal TT (>1/300) and no other US anomalies
Abnormal US screening suggestive for T21
T21 in previous history
Rob translocation 13, 21
Many other indications
NIPT optional, but not preferable
No indication for invasive…but NIPT desirable ??
Normal 1st trim and older age
Low risk 1st trim
Previous TT anomalies
ICSI /IVF/PGD pregnancies
…all pregnancies
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Practical information
Limitations of NIPT
Pregnancy duration
NIPT is less reliable if pregnancy < 12w
NIPT is impossible < 10 w
Maternal weight
NIPT is less reliable if high BMI
Weight (before pregnancy ) and length requested
Other factors influencing outcome
High physical activity
Infection
Cancer
…
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Practical information
Limitations of NIPT
Not detectable
Mosaïcism of chromosome 21
Deletions or duplications of chromosome 21
Other chromosomal anomalies
Molecular anomalies (monogenic ea CF, Fragile X…)
Misinterpretations can occur if
Maternal chromosomal anomalies or variants
Report on chromosomal testing of mother needed
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Practical information
Contra indications for NIPT
Twin or multiple pregnancy
History of (<3months)
maternal blood transfusion
stemcell therapy
immunotherapy
transplantation
US anomalies in foetus (
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NIPT - Non-invasive prenatal testing
array)
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Questions?
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NIPT - Non-invasive prenatal testing
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