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

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.

Evaluation of chronic lymphocytic leukemia by oligonucleotide-based microarray analysis uncovers novel aberrations not detected by FISH or cytogenetic analysis.

BACKGROUND: Cytogenetic evaluation is a key component of the diagnosis and prognosis of chronic lymphocytic leukemia (CLL). We performed oligonucleotide-based comparative genomic hybridization microarray analysis on 34 samples with CLL and known abnormal karyotypes previously determined by cytogenetics and/or fluorescence in situ hybridization (FISH). RESULTS: Using a custom designed microarray that targets >1800 genes involved in hematologic disease and other malignancies, we identified additional cryptic aberrations and novel findings in 59% of cases. These included gains and losses of genes associated with cell cycle regulation, apoptosis and susceptibility loci on 3p21.31, 5q35.2q35.3, 10q23.31q23.33, 11q22.3, and 22q11.23. CONCLUSIONS: Our results show that microarray analysis will detect known aberrations, including microscopic and cryptic alterations. In addition, novel genomic changes will be uncovered that may become important prognostic predictors or treatment targets for CLL in the future.

Effects of ozone exposure during microarray posthybridization washes and scanning.

The increasing prevalence of array-based comparative genomic hybridization in the clinical laboratory necessitates the implementation of quality control measures to attain accurate results with a high level of confidence. Environmental ozone is present in all industrialized cities and has been found to be detrimental to array data even at levels considered acceptable by US Environmental Protection Agency standards. In this study, we characterized the effect of ozone on microarray data on three different labeling platforms that use different fluorescent dyes (Cy3 and Cy5, Alexa Fluor 555 and Alexa Fluor 647, and Alexa Fluor 3 and Alexa Fluor 5) that are commonly used in array-based comparative genomic hybridization. We investigated the effects of ozone on microarray data by washing the array in variable ozone environments. In addition, we observed the effects of prolonged exposure to ozone on the microarray after washing in an ozone-free environment. Our results demonstrate the necessity of minimizing ozone exposure when washing and drying the microarray. We also found that washed microarrays produce the best results when immediately scanned; however, if a low-ozone environment is maintained, there will be little compromise in the data collected.