The tissue is the issue: improved methylome analysis from paraffin-embedded tissues by application of the HOPE technique

• Published in: Laboratory investigation 2014-08, Vol.94 (8), p.927-933
• Main Authors: Marwitz, Sebastian, Kolarova, Julia, Reck, Martin, Reinmuth, Niels, Kugler, Christian, Schädlich, Ines, Haake, Andrea, Zabel, Peter, Vollmer, Ekkehard, Siebert, Reiner, Goldmann, Torsten, Ammerpohl, Ole
• Format: Article
• Language: English
• Subjects: 631/1647/1407/1555; 631/1647/2210/2213; Adenocarcinoma - metabolism; Adenocarcinoma - pathology; Aged; Buffers; Carcinoma, Adenosquamous - metabolism; Carcinoma, Adenosquamous - pathology; Carcinoma, Large Cell - metabolism; Carcinoma, Large Cell - prevention & control; Carcinoma, Squamous Cell - metabolism; Carcinoma, Squamous Cell - pathology; Cell Line, Tumor; Cryopreservation; DNA Methylation; DNA, Neoplasm - isolation & purification; DNA, Neoplasm - metabolism; Female; Glutamic Acid - chemistry; HEPES - chemistry; Humans; Laboratory Medicine; Lung - metabolism; Lung - pathology; Lung Neoplasms - metabolism; Lung Neoplasms - pathology; Male; Medicine; Medicine & Public Health; Middle Aged; Pathology; technical-report; Tissue Array Analysis; Tissue Fixation; Tissue Preservation
• ISSN: 0023-6837; 1530-0307
• DOI: 10.1038/labinvest.2014.79

Alterations in the DNA methylome are characteristic for numerous diseases and a typical hallmark of cancer. Therefore, DNA methylation is currently under investigation in research labs and has also entered diagnostics. Recently, protocols like the BeadChip technology have become commercially available to study DNA methylation in an array format and semiquantitative fashion. However, it is known that fixation of the sample material with formalin prior to BeadChip analysis can affect the results. In this study we compared the influence of fixation on the outcome of BeadChip analysis. From six patients each a lung cancer tissue sample and a corresponding tumor-free lung tissue sample were collected. The samples were separated into three pieces. One piece of each sample was fixed with formalin, another one by the non-cross-linking HOPE technique (Hepes-glutamic acid buffer mediated Organic solvent Protection Effect). Subsequently, both became paraffin embedded. As a reference, the remaining third piece was cryopreserved. In addition we used three adenocarcinoma cell lines (H838, A549, and H1650) to validate the results from patient tissues. We show that using the HOPE technique instead of formalin largely prevents the introduction of formalin-fixation related artifacts. An ANOVA analysis significantly separated HOPE- and cryopreserved from formalin-fixed samples (FDR