New service: FFPE Methylation analysis
Illumina’s FFPE Restoration for Infinium experiments
Formalin-fixed paraffin-embedded (FFPE) tissue represents a valuable and extensive source of material for molecular analysis and retrospective clinical studies. However, the fixation and embedding procedures and subsequent storage affect the integrity of genomic DNA. DNA from FFPE samples is fragmented, chemically modified, depurinated and contains cross-links to other biomolecules. The challenge is to recover and be able to utilize the nucleic acids from embedded tissue samples, which may differ in the extent of the modification and damage to the DNA through fixation and embedding.
Degraded FFPE samples generally perform poorly with Illumina’s Infinium assay, as they are inefficiently amplified in the whole genome amplification step of the assay protocol. To overcome this problem, Illumina has developed the Infinium FFPE DNA Restore kit to repair damaged DNA and achieve high-quality genotyping, copy number variation (CNV) and methylation data using Infinium HD assays. During the restoration process, gaps and overhangs in the damaged double-stranded DNA are filled using the complement strand as a template.
In a pilot experiment at ServiceXS, the ability of the Illumina FFPE restoration kit to increase the quality of subsequent Infinium results clearly outperformed that of Qiagen’s REPLI-g ligation step, which was applied on FFPE material for Infinium Methylation analysis in a few recent publications (Thirlwell et al. (2010), Methods 52, 248-254; Jasmine et al. (2012), BMC Research Notes 5:117).
In the subsequent validation experiment, matching FFPE DNA and DNA from fresh frozen material was used in Infinium (methylation) experiments following restoration of the FFPE material, including bisulfite conversion prior to restoration. The results were very promising, with good reproducibility for replicate FFPE samples and a high number of detected CpG, combined with high concordance with fresh frozen (R2 >0.9) for 80% of restored samples passing entry QC. This opens up new possibilities for using DNA from FFPE material in customer projects at ServiceXS.
Contact us for more information on the possibilities for this new service. We are looking for EarlyXS customers.