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Publication

Facilitating the Fragile X post- and prenatal genetic diagnostic testing workflow by use of the Abbott FMR1 TP-PCR and FMR1 sizing PCR products.

Journal Contribution - Journal Article Conference Contribution

Molecular testing of FMR1 (CGG)n expanded repeats remains hard to tackle given the limitations of sizing PCR in detecting large and uninformative alleles, resulting in a significant number of samples that require confirmation through laborious southern blot analysis. Abbott recently developed a TP-PCR in combination with a sizing PCR to facilitate the Fragile X diagnostic testing workflow.
This study aimed at comparing Abbott FraXa sizing PCR and TP-PCR versus in-house sizing PCR and southern blot in order to evaluate the sizing accuracy and detection sensitivity of normal, intermediate, premutation and mutation alleles. Over 100 samples of different sources (approximately 50% whole blood, 40% chorion villi and 10% amniocytes) and a panel of artificially mimicked mosaic samples were evaluated.
Signal intensity and sizing accuracy met expectations in the normal to small premutation range with a deviation of ± 1 repeat regardless the sample source or DNA extraction method used. The sizing long run greatly improved the detection capacity and sizing accuracy of longer range (large premutation to full mutation) fragments, although inspection of raw data is recommended. TP-PCR allowed discrimination between normal/intermediate and premutation/mutation alleles. However, premutation and mutation TP-PCR signal patterns were very similar, therefore requiring the Abbott sizing PCR to distinguish them.
Mosaic detection in sizing- and TP-PCR ranged from 20% to 10% depending on the repeat range mixture used.
Our results corroborate the suggested Abbott workflow of using FraXa TP-PCR as a first line screening platform in combination with the sizing PCR as second line test.
Journal: Eur J Hum Genet
ISSN: 1018-4813
Volume: 19
Pages: 178-178
Publication year:2011
Keywords:Fragile X, FMR1, prenatal diagnosis