OC05.01: Automated isolation of single circulating fetal trophoblasts for prenatal genetic analysis

We present a novel multistep method to enrich and isolate circulating fetal extravillous trophoblasts (fEVTs) for molecular characterisation. The aims of this study were to determine the performances of the developed method, to assess the feasibility of downstream copy-number profiling on isolated single fEVTs, to evaluate the concordance rate with invasive diagnostic results and to define the resolution of our approach for copy-number variation (CNV) detection. fEVTs underwent enrichment using ferrofluid-conjugated antibodies in the CellTracks AutoPrep System. Individual fEVTs were isolated by DEPArray image-based system. Eventually, shallow sequencing of whole-genome amplified libraries using Ampli1 WGA and LowPass kits was performed for aneuploidies detection. Fetal identity was independently confirmed by means of genotyping and/or in-silico approach. Moreover, isolated single cells together with genomic DNA from the CNV Reference Panel (Coriell) were used to assess the workflow analytical sensitivity for microdeletion/duplication detection. A total of 372 women were enrolled to determine success rate and number of trophoblast per sample. At least one fetal cell was isolated in 90.7% of samples obtained at gestational age 10–11 weeks, and in 80.6% of samples obtained at 12–14 weeks. Among women with recoveries, the mean number of trophoblasts was 4.2 cells per patient at 10–11 weeks and 3.3 cells per patient at later weeks of gestation. Genetic analysis of fetal cells from 131 women resulted in identification of 15 cases with common chromosomal abnormalities, with 100% concordance with standard karyotyping following invasive diagnostic sampling. The analyses to evaluate the resolution of the lowpass WGS approach confirmed our capability to reproducibly detect CNVs down to 1.5Mb. This novel, fully automated, non-invasive method for isolation and testing of individual fetal cells could enable the routine detection of clinically relevant genomic alterations.