Piskorz, A.M. et al. (2016) Methanol-based fixation is superior to buffered formalin for next-generation sequencing of DNA from clinical cancer samples. Annals of Oncology, 27(3), pp. 532-539. (doi: 10.1093/annonc/mdv613) (PMID:26681675) (PMCID:PMC4769995)
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Abstract
Background Next-generation sequencing (NGS) of tumour samples is a critical component of personalised cancer treatment, but requires high quality DNA samples. Routine neutral-buffered formalin (NBF) fixation has detrimental effects on nucleic acids, causing low yields, as well as fragmentation and DNA base changes, leading to significant artefacts. Patients and Methods We have performed detailed comparison of DNA quality from matched samples isolated from high grade serous ovarian cancers from 16 patients fixed in methanol and NBF. These experiments use tumour fragments as well as mock biopsies to simulate routine practice, ensuring that results are applicable to standard clinical biopsies. Results Using matched snap frozen tissue as gold standard comparator, we show that methanol-based fixation has significant benefits over NBF, with greater DNA yield, longer fragment size and more accurate copy number calling using shallow whole genome sequencing. These data also provide a new approach to understand and quantify artefactual effects of fixation using non-negative matrix factorisation to analyse mutational spectra from targeted and whole genome sequencing data. Conclusion We strongly recommend the adoption of methanol fixation for sample collection strategies in new clinical trials. This approach is immediately available, is logistically simple and can offer cheaper and more reliable mutation calling than traditional NBF fixation.
Item Type: | Articles |
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Status: | Published |
Refereed: | Yes |
Glasgow Author(s) Enlighten ID: | Orange, Miss Clare and Ennis, Dr Darren and Mcneish, Professor Iain |
Authors: | Piskorz, A.M., Ennis, D., Macintyre, G., Goranova, T.E., Eldridge, M., Segui-Gracia, N., Valganon, M., Hoyle, A., Orange, C., Moore, L., Jimenez-Linan, M., Millan, D., Mcneish, I.A., and Brenton, J.D. |
College/School: | College of Medical Veterinary and Life Sciences > School of Cancer Sciences College of Medical Veterinary and Life Sciences > School of Medicine, Dentistry & Nursing |
Journal Name: | Annals of Oncology |
Publisher: | Oxford University Press |
ISSN: | 0923-7534 |
ISSN (Online): | 1569-8041 |
Copyright Holders: | Copyright © 2015 The Authors |
First Published: | First published in Annals of Oncology 27(3):532-539 |
Publisher Policy: | Reproduced under a Creative Commons License |
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