Abstract LB-178: Analysis of restored FFPE samples on Infinium methylation arrays

Abstract

Abstract Millions of Formalin Fixed Paraffin Embedded (FFPE) cancer tissue samples currently present in the US and worldwide provide an enormous, invaluable repository to the discovery of biomarkers in cancer research, drug development, diagnosis and treatment of diseases. However, the processes of fixation and storage of FFPE samples leads to degradation and base modification with chemical residues. This degraded FFPE DNA is inefficiently amplified by the Infinium® whole genome amplification (WGA) step leading to poor performance in genome wide methylation analysis. In this study, purified FFPE DNA or crude lysates (200–500 ng input) were bisulfite converted and restored through Illumina's Infinium HD DNA Restoration kit consisting of a combination of treatment with a DNA polymerase, a repair enzyme, and a ligase. Results of methylation analysis on high density HumanMethylation450 BeadChip®, featuring over 450,000 assays indicated that even highly degraded samples which show initial call rates (genes detected p =0.01) of 50–70% can be successfully restored to a state that achieved call rates of greater than 90%. Direct comparison of frozen and FFPE samples from the same tumors showed an R2 correlation of beta values of greater than 95.0% between fresh frozen (FF) and FFPE samples. We also developed a method to assess FFPE sample quality based on a 90 bp qPCR assay (Infinium HD FFPE QC kit). In this assay FFPE samples exhibiting a Ct (Cq) difference of less than six relative to an intact standard are considered to pass the QC test and generally perform well in the assay. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-178. doi:10.1158/1538-7445.AM2011-LB-178