Development and validation of an accurate quantitative real-time polymerase chain reaction–based assay for human blastocyst comprehensive chromosomal aneuploidy screening

A 4-hour quantitative real-time polymerase chain reaction method for preimplantation comprehensive 24-chromosome aneuploidy screening was developed and validated. Results demonstrated 97.6% accuracy on cell lines and 98.6% consistency with microarray-based blastocyst diagnoses.

Nathan R. Treff, Ph.D., Xin Tao, M.Sc., Kathleen M. Ferry, B.Sc., Jing Su, M.Sc., Deanne Taylor, Ph.D., Richard T. Scott Jr., M.D., H.C.L.D.
Volume 97, Issue 4 , Pages 819-824.e2, April 2012

To develop and validate a quantitative real-time polymerase chain reaction (qPCR)–based method for blastocyst trophectoderm comprehensive chromosome screening (CCS) of aneuploidy.

Prospective, randomized, and blinded.

Academic center for reproductive medicine.

Nine cell lines were obtained from a commercial cell line repository, and 71 discarded human blastocysts were obtained from 24 IVF patients that underwent preimplantation genetic screening.


Main Outcome Measure(s):
Consistency of qPCR diagnosis of aneuploidy compared with either conventional karyotyping of cell lines or microarray-based diagnoses of human blastocysts.

Samples from nine cell lines with well characterized karyotypes were diagnosed by qPCR with 97.6% (41/42) consistency. After applying a minimum threshold for concurrence, 100% consistency was achieved. Developmentally normal blastocysts designated as aneuploid or arrested blastocysts designated as euploid by single-nucleotide polymorphism microarray analyses were assigned identical 24 chromosome diagnoses by qPCR in 98.6% of cases (70/71). Overall euploidy (n = 37) and aneuploidy (n = 34) were assigned with 100% consistency. Data was obtained for both sample types in 4 hours.

These data demonstrate the first qPCR technology capable of accurate aneuploidy screening of all 24 chromosomes in 4 hours. This methodology provides an opportunity to evaluate trophectoderm biopsies with subsequent fresh euploid blastocyst transfer. Randomized controlled trials to investigate the clinical efficacy of qPCR-based CCS are currently underway.

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