Noninvasive prenatal exclusion of haemoglobin Bart's using foetal DNA from maternal plasma.
OBJECTIVE
Prenatal diagnosis of alpha-thalassaemia requires invasive testing associated with a risk of miscarriage. Cell-free foetal DNA in maternal plasma presents an alternative source of foetal genetic material for noninvasive prenatal diagnosis. We aimed to exclude HbBart's noninvasively by detection of unaffected paternal alleles in maternal plasma using quantitative fluorescence PCR (QF-PCR).


METHODS
Microsatellite markers (16PTEL05, 16PTEL06) within the breakpoint regions of -(SEA), -(FIL) and -(THAI) deletions were analysed using QF-PCR of maternal plasma from 30 families. In this blinded study, genotypes were confirmed using conventional PCR. Maternal plasma from two known cases of HbBart's were also analysed.


RESULTS
HbBart's was excluded in 10 out of 30 (33.3%, 95% CI, 17.3-52.8%) mothers by identifying the presence of nondeleted paternally inherited fetal alleles; either only 16PTEL05 (n = 1) or only 16PTEL06 (n = 4), or both (n = 5), and confirmed through direct analysis of fetal DNA. Paternally inherited foetal alleles of 16PTEL05 and 16PTEL06 were not detected in maternal plasma of the two known HbBarts cases. False negatives were excluded with the detection of paternally inherited fetal control marker, D21S1270 in maternal plasma.


CONCLUSIONS
We show proof-of-principle that such a test can accurately exclude HbBart's in the foetus by identifying the nondeleted paternally inherited fetal alleles in maternal plasma in one out of three pregnancies, avoiding invasive testing in these pregnancies.
Ho
Sherry S Y
SS
Department of Obstetrics and Gynaecology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore.
Chong
Samuel S C
SS
Koay
Evelyn S C
ES
Ponnusamy
Sukumar
S
Chiu
Lily
L
Chan
Yiong Huak
YH
Rauff
Mary
M
Baig
Sonia
S
Chan
Jerry
J
Su
Lin Lin
LL
Biswas
Arijit
A
Hahn
Sinuhe
S
Choolani
Mahesh
M
D000328:Adult; D004247:DNA / Q000235:genetics; D005220:Fathers; D005260:Female; D005453:Fluorescence; D017353:Gene Deletion; D006455:Hemoglobins, Abnormal / Q000032:analysis / Q000235:genetics*; D006801:Humans; D015160:Hydrops Fetalis / Q000097:blood / Q000175:diagnosis* / Q000235:genetics; D008297:Male; D008431:Maternal-Fetal Exchange; D018895:Microsatellite Repeats; D011237:Predictive Value of Tests; D011247:Pregnancy; D011296:Prenatal Diagnosis / Q000379:methods*; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D017085:alpha-Thalassemia / Q000097:blood / Q000175:diagnosis* / Q000235:genetics
D006455:Hemoglobins, Abnormal; D004247:DNA; C030448:hemoglobin Bart's