Comparison of array comparative genomic hybridization and quantitative real-time PCR-based aneuploidy screening of blastocyst biopsies

Abstract

Comprehensive chromosome screening (CCS) methods are being extensively used to select chromosomally normal embryos in human assisted reproduction. Some concerns related to the stage of analysis and which aneuploidy screening method to use still remain. In this study, the reliability of blastocyst-stage aneuploidy screening and the diagnostic performance of the two mostly used CCS methods (quantitative real-time PCR (qPCR) and array comparative genome hybridization (aCGH)) has been assessed. aCGH aneuploid blastocysts were rebiopsied, blinded, and evaluated by qPCR. Discordant cases were subsequently rebiopsied, blinded, and evaluated by single-nucleotide polymorphism (SNP) array-based CCS. Although 81.7% of embryos showed the same diagnosis when comparing aCGH and qPCR-based CCS, 18.3% (22/120) of embryos gave a discordant result for at least one chromosome. SNP array reanalysis showed that a discordance was reported in ten blastocysts for aCGH, mostly due to false positives, and in four cases for qPCR. The discordant aneuploidy call rate per chromosome was significantly higher for aCGH (5.7%) compared with qPCR (0.6%; P<0.01). To corroborate these findings, 39 embryos were simultaneously biopsied for aCGH and qPCR during blastocyst-stage aneuploidy screening cycles. 35 matched including all 21 euploid embryos. Blinded SNP analysis on rebiopsies of the four embryos matched qPCR. These findings demonstrate the high reliability of diagnosis performed at the blastocyst stage with the use of different CCS methods. However, the application of aCGH can be expected to result in a higher aneuploidy rate than other contemporary methods of CCS.

Figure 1

Flow chart of phase 1 preclinical validation of aCGH and qPCR-based aneuploidy screening of trophectoderm biopsies.

Figure 2

Profile plot comparison of some discordant cases between aCGH and qPCR with confirmation of diagnosis performed on a third biopsy and SNP array-based CCS. (a) False-positive and -negative chromosome error rates according to CCS methods under investigation. (b) Profile plots of aCGH showing a monosomy for the chromosome 18 for that embryo, shown to be normal by qPCR and SNP array-based CCS follow-up analysis. (c) aCGH profile plot consistent with a trisomy for chromosome 22 for that embryo, shown to be normal by qPCR and SNP array-based CCS follow-up analysis. (d) Profile plots of aCGH showing a double trisomy for chromosome 15 and 17 for that embryo, shown to be trisomy 15 by qPCR and SNP array-based CCS follow-up analysis.