Prenatal diagnosis of beta-thalassaemia using fetal erythroblasts enrichedfrom maternal blood by a novel gradient

We have assessed a new technique for the isolation of fetal erythroblasts f rom maternal blood for the noninvasive prenatal diagnosis of pregnancies at risk of beta-thalassaemia. This method relies on the separation of erythro blasts from maternal nucleated cells by a novel step gradient and high spee d centrifugation. In four of the six cases examined, single erythroblasts w ere identified by immunohistochemistry for zeta (zeta) globin. These were i ndividually micromanipulated and analysed by single cell polymerase chain r eaction (PCR) and subsequent sequencing of the region of beta-globin locus where the mutations most common to the region of Puglia, Italy, are cluster ed. In each of the four instances where fetal erythroblasts were identified by antibody staining, the fetal beta-globin genotype was correctly determi ned. To date, this represents the largest series of non-invasive prenatal d iagnoses performed for this haemoglobinopathy.