Evaluation of single‐nucleotide polymorphisms as internal controls in prenatal diagnosis of fetal blood groups. Issue 2 (13th June 2012)

• Record Type: Journal Article
• Authors: Doescher, Andrea; Petershofen, Eduard K.; Wagner, Franz F.; Schunter, Markus; Müller, Thomas H.
• Is Part Of: Transfusion. Volume 53:Issue 2(2013)
• Page Start: 353
• Page End: 362
• Publication Date: 2012-06-13
• Subjects: Hematology -- Periodicals; Blood -- Transfusion -- Periodicals; Blood Group Antigens -- Periodicals; Blood Preservation -- Periodicals; Blood Transfusion -- Periodicals; 615
• DOI: 10.1111/j.1537-2995.2012.03738.x
• Languages: English
• ISSNs: 0041-1132

Abstract

<abstract abstract-type="main" xml:lang="en"> <title> <x xml:space="preserve">Abstract</x> </title> <p> <bold>BACKGROUND:</bold> Determination of fetal blood groups in maternal plasma samples critically depends on adequate amplification of fetal DNA. We evaluated the routine inclusion of 52 single‐nucleotide polymorphisms (SNPs) as internal reference in our polymerase chain reaction (PCR) settings to obtain a positive internal control for fetal DNA.</p> <p> <bold>STUDY DESIGN AND METHODS:</bold> DNA from 223 plasma samples of pregnant women was screened for <italic>RHD</italic> Exons 3, 4, 5, and 7 in a multiplex PCR including 52 SNPs divided into four primer pools. Amplicons were analyzed by single‐base extension and the GeneScan method in a genetic analyzer. Results of D screening were compared to standard <italic>RHD</italic> genotyping of amniotic fluid or real‐time PCR of fetal DNA from maternal plasma.</p> <p> <bold>RESULTS:</bold> The vast majority of all samples (97.8%) demonstrated differences in maternal and fetal SNP patterns when tested with four primer pools. These differences were not observed in less than 2.2% of the samples most probably due to an extraction failure for adequate amounts of fetal DNA. Comparison of the fetal genotypes with independent results did not reveal a single false‐negative case among samples (n = 42) with positive internal control and negative fetal <italic>RHD</italic> typing.</p> <p> <bold>CONCLUSION:</bold> Coamplification of 52 SNPs with <italic>RHD</italic>‐specific sequences for fetal blood group determination introduces a valid positive control for the amplification of fetal DNA to avoid false‐negative results. This new approach does not require a paternal blood sample. It may also be applicable to other assays for fetal genotyping in maternal blood samples.</p> </abstract>