A multiplex PCR predictor for aCGH success of FFPE samples

Abstract

Formalin-fixed, paraffin-embedded (FFPE) tissue archives are the largest and longest time-spanning collections of patient material in pathology archives. Methods to disclose information with molecular techniques, such as array comparative genomic hybridisation (aCGH) have rapidly developed but are still not optimal. Array comparative genomic hybridisation is one efficient method for finding tumour suppressors and oncogenes in solid tumours, and also for classification of tumours. The fastest way of analysing large numbers of tumours is through the use of archival tissue samples with first, the huge advantage of larger median follow-up time of patients studied and second, the advantage of being able to locate and analyse multiple tumours, even across generations, from related individuals (families). Unfortunately, DNA from archival tissues is not always suitable for molecular analysis due to insufficient quality. Until now, this quality remained undefined. We report the optimisation of a genomic-DNA isolation procedure from FFPE pathology archives in combination with a subsequent multiplex PCR-based quality-control that simply identified all samples refractory to further DNA-based analyses. British Journal of Cancer (2006) Cancer cytogenetics has benefited greatly from the introduction of comparative genomic hybridisation (CGH) for mapping chromosomal gains and losses at a genome-wide scale (Kallioniemi et al, 1993;Gray et al, 1994). Subsequent development of the technique into array-CGH (also named matrix-CGH) has allowed increased automation, improved reproducibility and precision due to more accurate mapping of aberrations. This technology has been applied successfully to characterise congenital abnormalities at unprecedented precision (Veltman et al, 2002) and to characterise and classify tumours (Wessels et al, 2002;Nessling et al, 2005).In most pathology laboratories, large archives of formalin-fixed, paraffin embedded (FFPE) material are often the only source of material for cancer research. It is our experience (Wessels et al, 2002;Van Beers et al, 2005) that a proportion of archival specimens appears unsuited for aCGH analysis, which is troublesome because array comparative genomic hybridisation (aCGH) experiments are tedious and expensive. In the past, we have noticed that this was not solved by repeating aCGH experiments, even when DNA was isolated from new sections from the same tissue blocks (Van Beers et al, 2005). Nevertheless, it is possible to obtain high-quality data using archival DNA samples in array CGH experiments (Figure 1) (Gray et al, 1994;Ried et al, 1995;Albertson and Pinkel, 2003;Heidenblad et al, 2004;Loo et al, 2004;Devries et al, 2005), even from 20-year-old tissue blocks, provided that robust procedures, high-quality reagents and 'good' sample DNA quality are being used. A 'good sample quality' definition and an assay to determine this FFPE DNA sample quality would therefore be of great value.Molecular biological assays, including aCGH on FFPE archival specimens, would be more eff...

Exploring chromosomal abnormalities and genetic changes in uterine smooth muscle tumors

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Array CGH demonstrates characteristic aberration signatures in human papillary thyroid carcinomas governed by RET/PTC

“…To ensure that the extracted DNA was of required quality for use in array CGH experiments, a gene-specific multiplex PCR was used (van Beers et al, 2006). Only cases with amplification products larger or equal than 300 bp were designated to be of sufficient quality for array CGH analysis.…”

“…In this study, we used bacterial artificial chromosome (BAC)-arrays (B3400 clones) that allow the examination of the entire genome for chromosomal copy number changes (DNA gains and losses) with a resolution of 1 Mb. A striking advantage of this technology is its use on archival paraffinembedded material (van Beers et al, 2006).…”

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

“…For example, the introduction of a prequalifier PCR step provides a means of predicting which FFPE DNA samples might yield sufficient quality SNP array data. 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.…”