Evaluation of a novel assay for detection of the fetal marker RASSF1A: facilitating improved diagnostic reliability of noninvasive prenatal diagnosis.
<h4>Background</h4>Analysis of cell free fetal (cff) DNA in maternal plasma is used routinely for non invasive prenatal diagnosis (NIPD) of fetal sex determination, fetal rhesus D status and some single gene disorders. True positive results rely on detection of the fetal target being ana...
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Public Library of Science (PLoS)
2012-01-01
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| author | Helen E White Carolyn L Dent Victoria J Hall John A Crolla Lyn S Chitty |
| author_facet | Helen E White Carolyn L Dent Victoria J Hall John A Crolla Lyn S Chitty |
| author_sort | Helen E White |
| collection | DOAJ |
| description | <h4>Background</h4>Analysis of cell free fetal (cff) DNA in maternal plasma is used routinely for non invasive prenatal diagnosis (NIPD) of fetal sex determination, fetal rhesus D status and some single gene disorders. True positive results rely on detection of the fetal target being analysed. No amplification of the target may be interpreted either as a true negative result or a false negative result due to the absence or very low levels of cffDNA. The hypermethylated RASSF1A promoter has been reported as a universal fetal marker to confirm the presence of cffDNA. Using methylation-sensitive restriction enzymes hypomethylated maternal sequences are digested leaving hypermethylated fetal sequences detectable. Complete digestion of maternal sequences is required to eliminate false positive results.<h4>Methods</h4>cfDNA was extracted from maternal plasma (n = 90) and digested with methylation-sensitive and insensitive restriction enzymes. Analysis of RASSF1A, SRY and DYS14 was performed by real-time PCR.<h4>Results</h4>Hypermethylated RASSF1A was amplified for 79 samples (88%) indicating the presence of cffDNA. SRY real time PCR results and fetal sex at delivery were 100% accurate. Eleven samples (12%) had no detectable hypermethylated RASSF1A and 10 of these (91%) had gestational ages less than 7 weeks 2 days. Six of these samples were male at delivery, five had inconclusive results for SRY analysis and one sample had no amplifiable SRY.<h4>Conclusion</h4>Use of this assay for the detection of hypermethylated RASSF1A as a universal fetal marker has the potential to improve the diagnostic reliability of NIPD for fetal sex determination and single gene disorders. |
| format | Article |
| id | doaj-art-78beffdd43eb4363a6bad1bfbdb14f60 |
| institution | OA Journals |
| issn | 1932-6203 |
| language | English |
| publishDate | 2012-01-01 |
| publisher | Public Library of Science (PLoS) |
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| spelling | doaj-art-78beffdd43eb4363a6bad1bfbdb14f602025-08-20T02:30:56ZengPublic Library of Science (PLoS)PLoS ONE1932-62032012-01-0179e4507310.1371/journal.pone.0045073Evaluation of a novel assay for detection of the fetal marker RASSF1A: facilitating improved diagnostic reliability of noninvasive prenatal diagnosis.Helen E WhiteCarolyn L DentVictoria J HallJohn A CrollaLyn S Chitty<h4>Background</h4>Analysis of cell free fetal (cff) DNA in maternal plasma is used routinely for non invasive prenatal diagnosis (NIPD) of fetal sex determination, fetal rhesus D status and some single gene disorders. True positive results rely on detection of the fetal target being analysed. No amplification of the target may be interpreted either as a true negative result or a false negative result due to the absence or very low levels of cffDNA. The hypermethylated RASSF1A promoter has been reported as a universal fetal marker to confirm the presence of cffDNA. Using methylation-sensitive restriction enzymes hypomethylated maternal sequences are digested leaving hypermethylated fetal sequences detectable. Complete digestion of maternal sequences is required to eliminate false positive results.<h4>Methods</h4>cfDNA was extracted from maternal plasma (n = 90) and digested with methylation-sensitive and insensitive restriction enzymes. Analysis of RASSF1A, SRY and DYS14 was performed by real-time PCR.<h4>Results</h4>Hypermethylated RASSF1A was amplified for 79 samples (88%) indicating the presence of cffDNA. SRY real time PCR results and fetal sex at delivery were 100% accurate. Eleven samples (12%) had no detectable hypermethylated RASSF1A and 10 of these (91%) had gestational ages less than 7 weeks 2 days. Six of these samples were male at delivery, five had inconclusive results for SRY analysis and one sample had no amplifiable SRY.<h4>Conclusion</h4>Use of this assay for the detection of hypermethylated RASSF1A as a universal fetal marker has the potential to improve the diagnostic reliability of NIPD for fetal sex determination and single gene disorders.https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0045073&type=printable |
| spellingShingle | Helen E White Carolyn L Dent Victoria J Hall John A Crolla Lyn S Chitty Evaluation of a novel assay for detection of the fetal marker RASSF1A: facilitating improved diagnostic reliability of noninvasive prenatal diagnosis. PLoS ONE |
| title | Evaluation of a novel assay for detection of the fetal marker RASSF1A: facilitating improved diagnostic reliability of noninvasive prenatal diagnosis. |
| title_full | Evaluation of a novel assay for detection of the fetal marker RASSF1A: facilitating improved diagnostic reliability of noninvasive prenatal diagnosis. |
| title_fullStr | Evaluation of a novel assay for detection of the fetal marker RASSF1A: facilitating improved diagnostic reliability of noninvasive prenatal diagnosis. |
| title_full_unstemmed | Evaluation of a novel assay for detection of the fetal marker RASSF1A: facilitating improved diagnostic reliability of noninvasive prenatal diagnosis. |
| title_short | Evaluation of a novel assay for detection of the fetal marker RASSF1A: facilitating improved diagnostic reliability of noninvasive prenatal diagnosis. |
| title_sort | evaluation of a novel assay for detection of the fetal marker rassf1a facilitating improved diagnostic reliability of noninvasive prenatal diagnosis |
| url | https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0045073&type=printable |
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