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Abstract
Aims: Studies of DNA methylomes hold enormous promise for biomedicine but are hampered by the technological challenges of analyzing many samples cost-effectively. Recently, a major extension of the previous Infinium HumanMethylation27 BeadChip® (Illumina, Inc. CA, USA), called Infinium HumanMethylation450 (Infinium Methylation 450K; Illumina, Inc. CA, USA) was developed. This upgraded technology is a hybrid of two different chemical assays, the Infinium I and Infinium II assays, allowing (for 12 samples in parallel) assessment of the methylation status of more than 480,000 cytosines distributed over the whole genome. In this article, we evaluate Infinium Methylation 450K on cell lines and tissue samples, highlighting some of its advantages but also some of its limitations. In particular, we compare the methylation values of the Infinium I and Infinium II assays. Materials & methods: We used Infinium Methylation 450K to profile: first, the well-characterized HCT116 wild-type and double-knockout cell lines and then, 16 breast tissue samples (including eight normal and eight primary tumor samples). Absolute methylation values (β-values) were extracted with the GenomeStudio™ software and then subjected to detailed analysis. Results: While this technology appeared highly robust as previously shown, we noticed a divergence between the β-values retrieved from the type I and type II Infinium assays. Specifically, the β-values obtained from Infinium II probes were less accurate and reproducible than those obtained from Infinium I probes. This suggests that data from the type I and type II assays should be considered separately in any downstream bioinformatic analysis. To be able to deal with the Infinium I and Infinium II data together, we developed and tested a new correction technique, which we called ‘peak-based correction‘. The idea was to rescale the Infinium II data on the basis of the Infinium I data. While this technique should be viewed as an approximation method, it significantly improves the quality of Infinium II data. Conclusion: Infinium 450K is a powerful technique in terms of reagent costs, time of labor, sample throughput and coverage. It holds great promise for the better understanding of the epigenetic component in health and disease. Yet, due to the nature of its design comprising two different chemical assays, analysis of the whole set of data is not as easy as initially anticipated. Correction strategies, such as the peak-based approach proposed here, are a step towards adequate output data analysis.
Financial & competing interests disclosure
S Dedeurwaerder and M Defrance were supported by the Belgian ‘FNRS-Télévie‘ and ‘Interuniversity Attraction Poles‘ (IAP P6/28), respectively. E Calonne and H Denis were supported by the ULB and the Brussels Region ‘BruBreast‘. C Sotiriou and F Fuks are ‘Chercheur Qualifié‘ and ‘Maître de Recherche‘ from the FNRS This work was funded by grants from the FNRS and Télévie, the Brussels Region ‘BruBreast‘ and the ‘Interuniversity Attraction Poles‘ (IAP P6/28), by the EU grant CANCERDIP FP7-200620 and by a European Molecular Biology Organization Young Investigator Programme (EMBO YIP). The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.
No writing assistance was utilized in the production of this manuscript.
Ethical conduct of research
The authors state that they have obtained appropriate institutional review board approval or have followed the principles outlined in the Declaration of Helsinki for all human or animal experimental investigations. In addition, for investigations involving human subjects, informed consent has been obtained from the participants involved.