The development of a rapid assay for prenatal testing of common aneuploidies and microdeletion syndromes

Abstract

Objective To develop a novel, rapid prenatal assay for pregnancies with high likelihood of normal karyotypes, using BACs‐on‐Beads™ technology, a suspension array‐based multiplex assay that employs Luminex® xMAP® technology, for the detection of gains and losses in chromosomal DNA. Methods Fifteen relatively common microdeletions were selected that are not detectable, or may be missed, by karyotyping and usually do not present with abnormal ultrasound findings. Chromosomes 13, 18, 21, X, and Y were included. We validated the assay with 430 samples. Results All microdeletions and aneuploidies were correctly identified, except for a 69,XXX incorrectly identified as a normal female and a male with ∼20% maternal cell contamination (MCC) that could not be distinguished from 69,XXY. MCC became apparent at 20 to 30%. Mosaicism was identified at 30 to 35% abnormal cells. Conclusion We have developed an alternative to fluorescence in situ hybridization (FISH) aneuploidy screening and microarray analysis in otherwise normal pregnancies undergoing invasive testing. We demonstrated that the assay will detect all microdeletions and aneuploidies of regions covered on the assay. We developed analytical software that displays results for well‐characterized syndromes but not abnormalities of unclear clinical significance. This assay is likely to be preferred by women seeking testing beyond routine karyotyping but who desire more information than provided by aneuploidy FISH. Copyright © 2011 John Wiley & Sons, Ltd.

Prenatal BoBsTM in the cytogenetic analysis of products of spontaneous miscarriage

“…This rapid targeted assay is dependent on DNA extraction and does not require live or intact cells, and represents an interesting alternative to conventional cytogenetics that require cell culture. [5,9,11,12] In our study, cell culture failure prevented standard karyotype analysis of 38 samples. However, Prenatal BoBs TM provided a conclusive result in all cases, making it a useful tool for the cytogenetic analysis of POCs, especially when culture fails.…”

“…It has been shown that Prenatal BoBs TM can detect mosaicism in fetal tissue at a level of 20 -40% abnormal cells or higher, [9,15] and that MCC in the fetal tissue becomes apparent at a level of 20 -30% of normal female cells. [11] Prenatal BoBs has been described as more informative than rapid FISH and QF-PCR, which screen only frequent aneuploidies. [9] This technique also enabled us to provide rapid results with conclu sive outcomes within 24 hours of receipt of the sample, [5,9,11,12] which is about half the time of fast FISH aneuploidy screening, because it is possible to interpret the profiles of several samples in few minutes.…”

“…[11] Prenatal BoBs has been described as more informative than rapid FISH and QF-PCR, which screen only frequent aneuploidies. [9] This technique also enabled us to provide rapid results with conclu sive outcomes within 24 hours of receipt of the sample, [5,9,11,12] which is about half the time of fast FISH aneuploidy screening, because it is possible to interpret the profiles of several samples in few minutes. [5,9] Furthermore, it is cheaper than other technologies that are able to detect more alterations (array-CGH), being of the same order of cost as fast FISH.…”

“…[5] Prenatal BoBs TM is a targeted assay, so the loci of the genome that may have clinical relevance in an unbalanced state and that are not targeted by the probe set will go undetected. [5,9,11] Better coverage throughout the genome would lead to the detection of additional clinically relevant imbalances, but would also identify gains or losses of unknown or unclear clinical significance. [11,14] Moreover, Prenatal BoBs TM has some limitations in the detection of polyploidies (triploidies and tetraploidies), [5,9] which account for nearly 16% of abnormalities found by conventional karyotyping of POC.…”

Bacterial artificial chromosomes (BACs)-on-Beads™ as a diagnostic platform for the rapid aneuploidy screening of products of conception

“…Cytogenetic techniques that are not dependent on actively growing tissue are becoming more common, including quantitative fluorescence (QF)-PCR (2,3), multiplex ligation-dependent probe amplification (MLPA) (4,5), molecular karyotyping (6) and bacterial artificial chromosomes (BACs)-on-Beads™ (BoBs) (7)(8)(9)(10)(11). Of these techniques, BoBs offers the ability to assess small batch sizes at a relatively low cost with comprehensive coverage of all the chromosome arms, whilst complementing the highly interrogative array-based methods that are used in molecular karyotyping.…”