Fetal Sex and RHD Genotyping with Digital PCR Demonstrates Greater Sensitivity than Real-time PCR
dc.contributor.author | Sillence, KA | |
dc.contributor.author | Roberts, LA | |
dc.contributor.author | Hollands, HJ | |
dc.contributor.author | Thompson, HP | |
dc.contributor.author | Kiernan, Michele | |
dc.contributor.author | Madgett, TE | |
dc.contributor.author | Ross Welch, C | |
dc.contributor.author | Avent, Neil | |
dc.date.accessioned | 2015-11-11T17:58:28Z | |
dc.date.available | 2015-11-11T17:58:28Z | |
dc.date.issued | 2015-11-01 | |
dc.identifier.issn | 0009-9147 | |
dc.identifier.issn | 1530-8561 | |
dc.identifier.uri | http://hdl.handle.net/10026.1/3798 | |
dc.description.abstract |
<jats:title>Abstract</jats:title> <jats:sec> <jats:title>BACKGROUND</jats:title> <jats:p>Noninvasive genotyping of fetal RHD (Rh blood group, D antigen) can prevent the unnecessary administration of prophylactic anti-D to women carrying RHD-negative fetuses. We evaluated laboratory methods for such genotyping.</jats:p> </jats:sec> <jats:sec> <jats:title>METHODS</jats:title> <jats:p>Blood samples were collected in EDTA tubes and Streck® Cell-Free DNA™ blood collection tubes (Streck BCTs) from RHD-negative women (n = 46). Using Y-specific and RHD-specific targets, we investigated variation in the cell-free fetal DNA (cffDNA) fraction and determined the sensitivity achieved for optimal and suboptimal samples with a novel Droplet Digital™ PCR (ddPCR) platform compared with real-time quantitative PCR (qPCR).</jats:p> </jats:sec> <jats:sec> <jats:title>RESULTS</jats:title> <jats:p>The cffDNA fraction was significantly larger for samples collected in Streck BCTs compared with samples collected in EDTA tubes (P &lt; 0.001). In samples expressing optimal cffDNA fractions (≥4%), both qPCR and digital PCR (dPCR) showed 100% sensitivity for the TSPY1 (testis-specific protein, Y-linked 1) and RHD7 (RHD exon 7) assays. Although dPCR also had 100% sensitivity for RHD5 (RHD exon 5), qPCR had reduced sensitivity (83%) for this target. For samples expressing suboptimal cffDNA fractions (&lt;2%), dPCR achieved 100% sensitivity for all assays, whereas qPCR achieved 100% sensitivity only for the TSPY1 (multicopy target) assay.</jats:p> </jats:sec> <jats:sec> <jats:title>CONCLUSIONS</jats:title> <jats:p>qPCR was not found to be an effective tool for RHD genotyping in suboptimal samples (&lt;2% cffDNA). However, when testing the same suboptimal samples on the same day by dPCR, 100% sensitivity was achieved for both fetal sex determination and RHD genotyping. Use of dPCR for identification of fetal specific markers can reduce the occurrence of false-negative and inconclusive results, particularly when samples express high levels of background maternal cell-free DNA.</jats:p> </jats:sec> | |
dc.format.extent | 1399-1407 | |
dc.format.medium | Print-Electronic | |
dc.language | en | |
dc.language.iso | eng | |
dc.publisher | Oxford University Press (OUP) | |
dc.subject | Blood Specimen Collection | |
dc.subject | DNA | |
dc.subject | Female | |
dc.subject | Genotype | |
dc.subject | Genotyping Techniques | |
dc.subject | Humans | |
dc.subject | Male | |
dc.subject | Polymerase Chain Reaction | |
dc.subject | Pregnancy | |
dc.subject | Real-Time Polymerase Chain Reaction | |
dc.subject | Rh-Hr Blood-Group System | |
dc.subject | Sensitivity and Specificity | |
dc.subject | Sex Determination Analysis | |
dc.title | Fetal Sex and RHD Genotyping with Digital PCR Demonstrates Greater Sensitivity than Real-time PCR | |
dc.type | journal-article | |
dc.type | Evaluation Study | |
dc.type | Journal Article | |
dc.type | Research Support, Non-U.S. Gov't | |
plymouth.author-url | https://www.webofscience.com/api/gateway?GWVersion=2&SrcApp=PARTNER_APP&SrcAuth=LinksAMR&KeyUT=WOS:000364112200013&DestLinkType=FullRecord&DestApp=ALL_WOS&UsrCustomerID=11bb513d99f797142bcfeffcc58ea008 | |
plymouth.issue | 11 | |
plymouth.volume | 61 | |
plymouth.publication-status | Published | |
plymouth.journal | Clinical Chemistry | |
dc.identifier.doi | 10.1373/clinchem.2015.239137 | |
plymouth.organisational-group | /Plymouth | |
plymouth.organisational-group | /Plymouth/Faculty of Health | |
plymouth.organisational-group | /Plymouth/Faculty of Health/School of Biomedical Sciences | |
plymouth.organisational-group | /Plymouth/REF 2021 Researchers by UoA | |
plymouth.organisational-group | /Plymouth/REF 2021 Researchers by UoA/UoA01 Clinical Medicine | |
plymouth.organisational-group | /Plymouth/REF 2021 Researchers by UoA/UoA01 Clinical Medicine/UoA01 Clinical Medicine | |
plymouth.organisational-group | /Plymouth/Research Groups | |
plymouth.organisational-group | /Plymouth/Research Groups/Institute of Translational and Stratified Medicine (ITSMED) | |
plymouth.organisational-group | /Plymouth/Research Groups/Institute of Translational and Stratified Medicine (ITSMED)/CBR | |
plymouth.organisational-group | /Plymouth/Research Groups/Plymouth Institute of Health and Care Research (PIHR) | |
plymouth.organisational-group | /Plymouth/Users by role | |
plymouth.organisational-group | /Plymouth/Users by role/Academics | |
dc.publisher.place | England | |
dcterms.dateAccepted | 2015-08-24 | |
dc.identifier.eissn | 1530-8561 | |
dc.rights.embargoperiod | Not known | |
rioxxterms.versionofrecord | 10.1373/clinchem.2015.239137 | |
rioxxterms.licenseref.uri | http://www.rioxx.net/licenses/all-rights-reserved | |
rioxxterms.licenseref.startdate | 2015-11 | |
rioxxterms.type | Journal Article/Review |