CGH arrays compared for DNA isolated from formalin‐fixed, paraffin‐embedded material

Abstract

Formalin-fixed, paraffin-embedded (FFPE) archival tissue is an important source of DNA material. The most commonly used technique to identify copy number aberrations from chromosomal DNA in tumorigenesis is array comparative genomic hybridization (aCGH). Although copy number analysis using DNA from FFPE archival tissue is challenging, several research groups have reported high quality and reproducible DNA copy number results using aCGH. Aim of this study is to compare the commercially available aCGH platforms suitable for high-resolution copy number analysis using FFPE-derived DNA. Two dual channel aCGH platforms (Agilent and NimbleGen) and a single channel SNP-based platform (Affymetrix) were evaluated using seven FFPE colon cancer samples, and median absolute deviation (MAD), deflection, signal-to-noise ratio, and DNA input requirements were used as quality criteria. Large differences were observed between platforms; Agilent and NimbleGen showed better MAD values (0.13 for both) compared with Affymetrix (0.22). On the contrary, Affymetrix showed a better deflection of 0.94, followed by 0.71 for Agilent and 0.51 for NimbleGen. This resulted in signal-to-nose ratios that were comparable between the three commercially available platforms. Interestingly, DNA input amounts from FFPE material lower than recommended still yielded high quality profiles on all platforms. Copy number analysis using DNA derived from FFPE archival material is feasible using all three high-resolution copy number platforms and shows reproducible results, also with DNA input amounts lower than recommended.

Evaluating the repair of DNA derived from formalin-fixed paraffin-embedded tissues prior to genomic profiling by SNP–CGH analysis

“…The restoration protocol required 100 ng of double-stranded DNA, which is less than the recommended 250 ng or more required for the Agilent or Nimblegen assays. Reduced DNA input amounts have been tested using Agilent, Nimblegen, and Affymetrix OncoScan FFPE platforms, 24 and these assays do show tolerance for lower input amounts (50-100 ng), but there is also some increase in data noise.…”

“…15,20,22 The recently developed Oncoscan FFPE platform (Affymetrix), a high-resolution SNP array based on molecular inversion probes, appears to perform well in comparison with aCGH platforms and so might prove to be a successful method for studying FFPE samples for CNAs. 23,24 Finally, there are now several commercial methods available designed to repair DNA that is damaged by tissue processing or sample storage. In these, overhangs and gaps in degraded DNA and adducts in chemically modified DNA sequences are enzymatically repaired, and short DNA fragments are ligated to produce longer fragments.…”

Whole genome profiling and other high throughput technologies in lymphoid neoplasms—current contributions and future hopes

“…These platforms also allow the use of DNA extracted from routinely processed formalin-fixed and paraffin-embedded (FFPE) tissues. 5 CGH arrays have an increased sensitivity, have a more homogeneous coverage of the genome and may detect small alterations with higher resolution but they need the combination of a normal reference DNA with the tumor DNA. The use of the constitutional DNA of the same patient provides the advantage of filtering out the individual CNV.…”

“…However, recent improvements in protocols that use nucleic acid extracted from FFPE routinely processed tissues for microarrays increases the likelihood that these technologies can be used in routine practice. 5,78 The information generated up to now has been based on the use of frozen samples. Validation studies using these new protocols for routine samples will be necessary to confirm the applicability of the results.…”

Lymphoma classification and the tools of our trade: an introduction to the 2012 USCAP Long Course

“…47 And more than just on the horizon in terms of clinical testing are the already well-established high-throughput techniques of array comparative genomic hybridization (aCGH) to detect copy number variations and single-nucleotide polymorphism (SNP) arrays to detect copy number variations and uniparental disomies (Figure 9). 49,50 One may also be able to suggest the possible presence of translocations; 51 however, at least currently, this is not the major strength or purpose of these techniques. As indicated above, coming along right behind, and even before aCGH and SNP arrays have been incorporated in many clinical practices, is next-generation sequencing.…”