Detection of Chromosome Aneuploidies in Chorionic Villus Samples by Multiplex Ligation-Dependent Probe Amplification

Abstract

The objective of this study was to examine the suitability of multiplex ligation-dependent probe amplification (MLPA) in chorionic villus samples as a replacement for traditional karyotyping for the detection of (an)euploidies of chromosomes 21 , 18 , 13, X, and Y. Chorionic villus samples were diagnosed by traditional karyotyping using short-term cultures (STC) and long-term cultures (LTC) , and by MLPA using kit P095. DNA was extracted after digestion of whole villi with proteinase K and/or trypsin and collagenase. Different cell-dissociation procedures were tested to obtain MLPA results representative of the cytotrophoblast layer and the mesenchymal core. Over 95% of the MLPA results were in concordance with the traditional karyotyping of STC and LTC. Traditional karyotyping revealed seven mosaics. After digestion of whole villi with proteinase K , only abnormal cell lines confined to the STC gave rise to abnormal MLPA results. In one sample , the complete discrepancy between STC and LTC was resolved after enzymatic dissociation of cells from the cytotrophoblast layer and the mesenchymal core. MLPA in chorionic villus samples was found to be a reliable test for the detection of (an)euploidies of chromosomes 21, 18 , 13 , X , and Y. Whole villi digestion with proteinase K resulted in the over-representation of cytotrophoblasts in the DNA pool. To obtain MLPA results representative for STC and LTC , enzymatic dissociation of cells from the cytotrophoblast layer and mesenchymal core is

Utility and limitations of multiplex ligation-dependent probe amplification technique in the detection of cytogenetic abnormalities in products of conception

“…[9] It can also not detect low-level mosaicism, maternal cell contamination, any interstitial segment copy number gain or loss located elsewhere in the genome other than subtelomeric region. [10] aCGH can detect copy number gains or loss located anywhere all over the genome, but using it to detect numerical abnormality in abortus sample will be very expensive. [4] Both MLPA and aCGH cannot detect balanced translocation.…”

Is routine karyotyping required in prenatal samples with a molecular or metabolic referral?

“…With the application of a cell dissociation protocol steps are taken to prevent fully discrepant results between cytotrophoblasts and mesenchymal core cells for the chromosomes 13, 18, 21, X and Y. Herewith, the cytotrophoblast cells (analyzed in STC) and mesenchymal core cells (analyzed in LTC) can be tested for RAD separately [29,30] and, thus, discrepancies as in cases 8 and 10 would have been detected if RAD were used. The application of RAD to replace karyotyping in both STC and LTC will prevent findings without clinical relevance and/or uncertain outcome.…”

Evaluation of Quantitative Fluorescence in situ Hybridization for Relative Measurement of Telomere Length in Placental Mesenchymal Core Cells

“…These cells had undergone a double enzymatic treatment (trypsin then collagenase) identical to that used before culture of the placental villi. Enzymatic dissociation with trypsin followed by collagenase has been shown to isolate cell populations consisting predominantly of cells from the mesenchymal core [23,24,25,26,27]. In the uncultured cells, the distribution of fluorescence intensity was normal (fig.…”