Evaluation of the clinical usefulness of isolation of fetal DNA from the maternal circulation

Objective: To assess the reliability of isolating free fetal DNA from maternal plasma and to evaluate its clinical usefulness. Design: Fetal DNA was isolated from plasma or serum that was either collected prospectively or from archived samples collected for the purposes of second trimester screening. Methods: Prospective samples were collected from patients undergoing prenatal diagnostic procedures (n = 24). A second group of samples from Rhesus negative women (n = 28) were assayed in which blood had originally been collected for maternal triple serum screening. DNA was extracted from all samples and assayed for the presence of the beta-globin gene, sex-determining region Y (SRY) gene and Rh gene. All DNA sample handling and extraction was carried out by a single operator, and polymerase chain reaction (PCR) was carried out using previously published PCR primers and appropriate controls. The accuracy of results was assessed relative to the karyotype in the case of the SRY gene or cord blood phenotype in the case of the Rh gene. Results: The SRY PCR results were compared to fetal cell karyotypes obtained from invasive diagnostic testing, 21 out of the 24 samples were correctly 'sexed'. The RhD PCR results were compared to fetal cord blood samples at the time of delivery, and showed both false positive and false negative results. Two RhD negative babies were genotyped as RhD positive, despite repeat analysis. Conclusion: It is possible to isolate fetal DNA from maternal serum. It is a potentially clinically useful technique in our laboratory and can be used to detect male fetuses, and Rh negative fetuses. To be useful in clinical practice, it is necessary to safeguard against contamination at the time of sample handling, and to use the optimal range of primers available to cover the polymorphisms present within the RhD gene. Although not robust enough yet to be used with diagnostic certainty in our hands, immense improvements in technique, probes and real-time PCR equipment make this type of diagnosis a reality in the near future.