Open AccessComparison of PCR methods for detecting fetal RhDin maternal plasma
Author(s) -
Atamaniuk Johanna,
Stuhlmeier Karl M.,
Karimi Alireza,
Mueller Mathias M.
Publication year - 2009
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/jcla.20282
Subject(s) - taqman , gestation , cell free fetal dna , fetus , andrology , real time polymerase chain reaction , pregnancy , melting curve analysis , gestational age , medicine , biology , microbiology and biotechnology , prenatal diagnosis , genetics , gene
Background : Aim of this study was to establish the method yielding the highest sensitivity routinely used to determine fetal RhD type and gender from maternal cell‐free plasma DNA in different periods of gestation. Methods : Plasma DNA concentrations were measured from 46 pregnant women in different gestational periods and tested for RhD using three different PCR methods on exon 7: Thermal Cycler, Taqman method on LightCycler, and melting curve analysis on LightCycler. In addition, fetal gender was determined by PCR. Cell‐free plasma DNA was measured in 100 healthy volunteers as a reference group. Results : The mean value of cell‐free plasma DNA in the reference group was 10.9 pg/µL mean, (standard deviation (SD): 3.66) in 50 healthy women and 12.7 pg/µL (SD: 8.2) in 50 healthy men. In the firsttrimester of pregnancy cell‐free plasmaDNA was 14.9 pg/µL mean, (SD: 4.2), in the second trimester 15.4 pg/µL mean, (SD: 4.96), and the maximum was achieved in the third trimester of pregnancy 15.6 pg/µl mean, (SD: 6.49). TaqMan probes had the same accuracy, when compared with Thermal Cycler technology (46 samples, 6 failures). Using real‐time PCR with melting curve analysis 12 of 17 samples were correctly tested. Gender determination was correctly in 41 of 46 samples. Conclusion : RhD determinations with TaqMan and Thermal Cycler technology are useful methods for fetal RhD prediction. To increase the accuracy of RhD determination it is necessary to test on other exons in addition. J. Clin. Lab. Anal. 23:24–28, 2009. © 2009 Wiley‐Liss, Inc.