1. Prenatal sampling and considerations

Question 1

What is ffDNA?

Where does it originate from?

Answer 1

Fetal DNA circulating in maternal blood (2-6%)

Apoptotic trophoblasts in placenta

Question 2

How do ffDNA levels change during pregnancy?

Answer 2

Present from 5 weeks, enough for testing at 10 weeks

Increases as pregnancy progresses

Question 3

How is ffDNA distinguished from maternal free DNA?

Answer 3

Shorter fragments, approximately 200bp

Question 4

What is a CVS? What is it composed of?

Answer 4

Chorionic villi are part of placental tissue

Composed of cytotrophoblast and mesenchyme core

Question 5

What quantity of CVS material is needed for testing?

Answer 5

Generally 5-10mg need cultured cells for DNA extraction

> 10mg - DNA extracted from uncultured cells

Question 6

What is the risk associated with a CVS?

Answer 6

Invasive procedure, 1-2% risk of miscarriage

Question 7

At what gestation is a CVS offered?

Answer 7

11-13 weeks (1st trimester) - earlier than amnio

Question 8

When is amniocentesis offered?

Answer 8

From 15 weeks - 2nd or 3rd trimester

Question 9

What is the risk associated with amniocentesis?

Answer 9

0.5-1% risk of miscarriage

Question 10

Why does AF give a better representation of the fetal karyotype than CVS?

Answer 10

Cell population in AF is heterogenous - derived from fetal tissue, extra-embryonic membranes, amniocytes from amnion

Question 11

When is fetal blood sampling used?

What is the advantage & disadvantage?

Answer 11

Following ambiguous results of CVS/amnio

Reliable indicator of fetal karyotype but higher risk of miscarriage

Question 12

What causes MCC in a CVS and an amnio

Answer 12

CVS - maternal decidua
Amnio - blood staining

Question 13

How does culturing affect MCC for AF and CVS?

Answer 13

AF - MCC lower in cultures than uncultured - culture conditions favour growth of amniocytes over maternal blood cells

CVS - MCC higher in cultures than uncultured as both cell types are cultured

Question 14

How is MCC tested for?

Answer 14

QF-PCR for microsatellite markers for 13, 18, 21, X, Y

Question 15

Why must MCC be excluded for all single gene tests?

Answer 15

MCC may compromise validity of results

False positive or negative is maternal DNA is tested

MLPA and TP-PCR are particular sensitive to MCC

Question 16

What is confined placental mosaicism?

Answer 16

Presence of abnormal cells in in extraembryonic tissues (placenta), not in fetus

Question 17

What cell types is CPM found in?

How often is CPM found?

Answer 17

50% trophoblasts
30% mesenchyme
20% both

Found in 1-2% of pregnancies at 10-12 weeks

Question 18

What is the most common abnormality found in CPM?

Answer 18

Trisomic line in placenta, normal diploid complement in fetus

Question 18

What are the two types of CPM? What cause them?

Answer 18

1. Mitotic CPM - mitotic non-disjunction in trophoblast cells –> trisomy 2, 3, 7, 8
2. Meiotic CPM - trisomic conception rescued in cells destined to become fetus, remaining trisomic cells confined to placenta –> trisomy 16, 22

Question 19

How does the location of CPM relate to the cause?

Answer 19

CPM in trophoblasts or mesenchyme = mitotic non-disjunction

CPM in both trophoblasts and mesenchyme = Trisomy rescue following meiotic non-disjunction

Question 19

What risk is associated with CPM caused by trisomy rescue?

Answer 19

UPD in ‘rescued’ diploid fetus - both chromosomes may be from same parent

If chr 6, 7, 11, 14, 15, 20 are involved

Question 19

What does aneuploidy on CVS but normal fetal karyotype on AF suggest?

Answer 19

CPM

Therefore risk of UPD/imprinting defect in fetus if chr 6, 7, 11, 14, 15, 20 are involved due to trisomy rescue

Question 20

Which cells in a CVS are most representative of the fetal karyotype?

Answer 20

Mesenchyme core - has similar embryological origin