CVS analysis by QF-PCR: advantages and limitations from an experience of 41197 first-trimester cytogenetic diagnoses
Francesca R. Grati1, Francesca Malvestiti1, Livia Marcato1, Beatrice Grimi1, Elisa Gaetani1, Anna Maria Di Meco1, Anna Trotta1, Rosaria Liuti1, Sara Chinetti1, Francesca Dulcetti1, Anna Maria Ruggeri1, Cristina Agrati1, Lorenza Martinoni1, Silvia Paganini, Barbara Castoldi, Beatrice Negri, Federico Maggi1, Giuseppe Simoni1
1Research and Development, Cytogenetics and Molecular Biology, TOMA Advanced Biomedical Assays, Busto Arsizio (VA), Italy
Objectives Karyotyping on chorionic villous samples (CVS) includes cytogenetic analysis of both cytotrophoblast (short term cultures, STC) and mesenchyme (long term cultures, LTC). This approach requires complex laboratory organization, standardization of cytogenetic methods and highly trained operators. This approach has the highest reliability and diagnostic yield in all invasive prenatal diagnosis indications. The introduction of QF-PCR instead of the conventional karyotype in low risk pregnancies opened suggestions about its application also in CVS analysis. Due to the feto-placental mosaicisms, discordant QF-PCR and CVS cytogenetic results were reported and strategies for CVS analysis were introduced to minimize these risks. Recently, the possibility to substitute the STC with QF-PCR has been reported. This approach (QF-PCR+LTC) simplifies and improve the rapidity of the diagnostic processes. The aim of this study is to evaluate benefits and limitations of the approach QF-PCR+LTC compared to the traditional method STC+LTC and to quantify the associated risks of false results.
Method This study is based on a large retrospective cytogenetic CVS datasets (n=41197) generated by STC+LTC analytic approach. Basing on these results the false negative risks related to TFM-IV cases, imprinting syndromes and maternal cell contamination in LTC are calculated. A comparison of the two methods in terms of confirmation on amniocytes (AF), costs evaluation, turnaround time (TAT) and staff training is also performed.
Results Compared to STC+LTC method the QF-PCR+LTC approach is associated with a cumulative false negative risk of ~1/2000. Costs and TAT of STC in an horizontally organized lab are similar to a CE-IVD marked QF-PCR analysis. The required staff training period in cytogenetics is 5-times higher than for QF-PCR.
Conclusions QF-PCR in substitution of STC for CVS analysis is associated a with false negative risk of about 1/2000. The only element favourable to STC substitution is the required staff training for direct preparation cytogenetic analysis.
Learning objective:
Participants shall be able to evaluate benefits, limitations and associated risks of false results of the approach “QF-PCR + long term culture” compared to the traditional method “short term culture + long term culture” for the cytogenetic analysis of chrionic villi
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