Optimal detection of fetal chromosomal abnormalities by massively parallel DNA sequencing of cell-free fetal DNA from maternal blood

cell-free fetal DNA from maternal blood can detect fe-tal chromosomal abnormalities. Although existing al-gorithms focus on the detection of fetal trisomy 21 (T21), these same algorithms have difficulty detecting trisomy 18 (T18). METHODS: Blood samples were collected from 1014 pa-tients at 13 US clinic locations before they underwent an invasive prenatal procedure. All samples were pro-cessed to plasma, and the DNA extracted from 119 samples underwent massively parallel DNA sequenc-ing. Fifty-three sequenced samples came from women with an abnormal fetal karyotype. To minimize the intra- and interrun sequencing variation, we developed an optimized algorithm by using normalized chromo-some values (NCVs) from the sequencing data on a training set of 71 samples with 26 abnormal karyo-types. The classification process was then evaluated on an independent test set of 48 samples with 27 abnormal karyotypes. RESULTS: Mapped sites for chromosomes of interest in the sequencing data from the training set were normal-ized individually by calculating the ratio of the number of sites on the specified chromosome to the number of sites observed on an optimized normalizing chromo-some (or chromosome set). Threshold values for tri-somy or sex chromosome classification were then es-tablished for all chromosomes of interest, and a classification schema was defined. Sequencing of the independent test set led to 100 % correct classification of T21 (13 of 13) and T18 (8 of 8) samples. Other chro-mosomal abnormalities were also identified. CONCLUSION: Massively parallel sequencing is capable of detecting multiple fetal chromosomal abnormalities frommaternal plasma when an optimized algorithm is used