OBJECTIVE: To examine the accuracy of fetal RHD genotype and RHD pseudogene determination in a multiethnical population. METHODS: Prospective study involving D-negative pregnant women. Cell-free DNA was extracted from 1 ml of maternal plasma by an automated system (MagNA Pure Compact, Roche) and real-time PCR was performed in triplicate targeting the RHD gene exons 5 and 7. Inconclusive samples underwent RHD pseudogene testing by real-time PCR analysis employing novel primers and probe. RESULTS: A positive result was observed in 128/185 (69.2%) samples and negative in 50 (27.0%). Umbilical cord blood phenotype confirmed all cases with a positive or negative PCR result. Seven (3.8%) cases were found inconclusive (exon 7 amplification only) and RHD pseudogene testing with both conventional and real-time PCR demonstrated a positive result in five of them, while two samples were also RHD pseudogene negative. CONCLUSION: Real-time PCR targeting RHD exons 5 and 7 simultaneously in maternal plasma is an accurate method for the diagnosis of fetal D genotype in our population. The RHD pseudogene real-time PCR assay is feasible and is particularly useful in populations with a high prevalence of this allele.
OBJECTIVE: To examine the accuracy of fetal RHD genotype and RHD pseudogene determination in a multiethnical population. METHODS: Prospective study involving D-negative pregnant women. Cell-free DNA was extracted from 1 ml of maternal plasma by an automated system (MagNA Pure Compact, Roche) and real-time PCR was performed in triplicate targeting the RHD gene exons 5 and 7. Inconclusive samples underwent RHD pseudogene testing by real-time PCR analysis employing novel primers and probe. RESULTS: A positive result was observed in 128/185 (69.2%) samples and negative in 50 (27.0%). Umbilical cord blood phenotype confirmed all cases with a positive or negative PCR result. Seven (3.8%) cases were found inconclusive (exon 7 amplification only) and RHD pseudogene testing with both conventional and real-time PCR demonstrated a positive result in five of them, while two samples were also RHD pseudogene negative. CONCLUSION: Real-time PCR targeting RHD exons 5 and 7 simultaneously in maternal plasma is an accurate method for the diagnosis of fetal D genotype in our population. The RHD pseudogene real-time PCR assay is feasible and is particularly useful in populations with a high prevalence of this allele.
Authors: Sina P Müller; Iris Bartels; Werner Stein; Günther Emons; Kai Gutensohn; Michael Köhler; Tobias J Legler Journal: Transfusion Date: 2008-08-07 Impact factor: 3.157
Authors: C Ellen van der Schoot; G H Martine Tax; Robbert J P Rijnders; Masja de Haas; Godelieve C M L Christiaens Journal: Transfus Med Rev Date: 2003-01
Authors: R J P Rijnders; R B Van Der Luijt; E D J Peters; J K Goeree; C E Van Der Schoot; J K Ploos Van Amstel; G C M L Christiaens Journal: Prenat Diagn Date: 2003-12-30 Impact factor: 3.050
Authors: Tobias J Legler; Zhong Liu; Ariadni Mavrou; Kirstin Finning; Ilona Hromadnikova; Silvia Galbiati; Cathy Meaney; Maj A Hultén; Francesco Crea; Martin L Olsson; Deborah G Maddocks; Dorothy Huang; Sylvia Armstrong Fisher; Markus Sprenger-Haussels; Aicha Ait Soussan; C Ellen van der Schoot Journal: Prenat Diagn Date: 2007-09 Impact factor: 3.050