Noninvasive Prenatal Diagnosis of Beta-Thalassemia Disease by Using Digital PCR Analysis of Cell-Free Fetal DNA in Maternal Plasma

Abstract

<b><i>Introduction:</i></b> Prenatal diagnosis of thalassemia disease was usually based on invasive technique<b>.</b> Noninvasive diagnosis using cell-free fetal DNA (cff-DNA) was described with various laboratory techniques. The aim of this study was to identify the performance of dPCR for analyzing cff-DNA in maternal plasma to diagnose fetal beta-thalassemia diseases. <b><i>Methods:</i></b> Thirty-five couples at risk of fetal beta-thalassemia disease caused by four common mutations of <i>HBB</i> were enrolled at 12–18 weeks. The dPCR assay was designed to detect and quantify paternally inherited beta-thalassemia allele (PIB) and maternally inherited beta-thalassemia allele (MIB) from cff-DNA in maternal plasma. <b><i>Results:</i></b> Of 29 couples with different paternal/maternal mutations, all cases who inherited paternal mutation had detectable PIB-M. The MIB-mutant/wild-type (MIB-M/MIB-N) ratio in the mothers whose fetuses did not inherit maternal mutation was 0.87 ± 0.07 which was significantly lower than that of the mothers whose fetuses inherited maternal mutation, 1.01 ± 0.05. The sensitivity and specificity of MIB-M/MIB-N ratio &#x3e;0.95 in predicting fetus inheriting maternal mutation were 100 and 92.3%, respectively. In four couples with same paternal/maternal mutation, IB-M/IB-N ratio of &#x3e;0.95 correctly predicted the presence of an inheritance of at least one beta-thalassemia allele. In two couples with paternal Hb E/beta-thalassemia, the presence of PIB-M and the MIB-M/MIB-N ratio of &#x3e;0.95 correctly predicted the presence of paternal/maternal mutations, respectively. <b><i>Conclusions:</i></b> The method of analyzing cff-DNA in maternal plasma by dPCR is efficient for prenatal diagnosis of beta-thalassemia.