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Reproducibility of molecular phenotypes after long-term differentiation to uuman iPSC-derived neurons: a multi-site omics study

Volpato, Viola, Smith, James, Sandor, Cynthia, Ried, Janina S., Baud, Anna, Handel, Adam, Newey, Sarah E., Wessely, Frank, Attar, Moustafa, Whiteley, Emma, Chintawar, Satyan, Verheyen, An, Barta, Thomas, Lako, Majlinda, Armstrong, Lyle, Muschet, Caroline, Artati, Aanna, Cusulin, Carlo, Christensen, Klaus, Patsch, Christoph, Sharma, Eshita, Nicod, Jerome, Brownjohn, Philip, Stubbs, Victoria, Heywood, Wendy E., Gissen, Paul, De Filippis, Roberta, Janssen, Katharina, Reinhardt, Peter, Adamski, Jerzy, Royaux, Ines, Peeters, Pieter J., Terstappen, Georg C., Graf, Martin, Livesey, Frederick J., Akerman, Colin J., Mills, Kevin, Bowden, Rory, Nicholson, George, Webber, Caleb, Cader, M. Zameel and Lakics, Viktor 2018. Reproducibility of molecular phenotypes after long-term differentiation to uuman iPSC-derived neurons: a multi-site omics study. Stem Cell Reports 11 (4) , pp. 897-911. 10.1016/j.stemcr.2018.08.013

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Abstract

Reproducibility in molecular and cellular studies is fundamental to scientific discovery. To establish the reproducibility of a well-defined long-term neuronal differentiation protocol, we repeated the cellular and molecular comparison of the same two iPSC lines across five distinct laboratories. Despite uncovering acceptable variability within individual laboratories, we detect poor cross-site reproducibility of the differential gene expression signature between these two lines. Factor analysis identifies the laboratory as the largest source of variation along with several variation-inflating confounders such as passaging effects and progenitor storage. Single-cell transcriptomics shows substantial cellular heterogeneity underlying inter-laboratory variability and being responsible for biases in differential gene expression inference. Factor analysis-based normalization of the combined dataset can remove the nuisance technical effects, enabling the execution of robust hypothesis-generating studies. Our study shows that multi-center collaborations can expose systematic biases and identify critical factors to be standardized when publishing novel protocols, contributing to increased cross-site reproducibility.

Item Type: Article
Date Type: Published Online
Status: Published
Schools: Medicine
Publisher: Elsevier
ISSN: 2213-6711
Date of First Compliant Deposit: 21 October 2020
Date of Acceptance: 21 August 2018
Last Modified: 21 Oct 2020 21:07
URI: http://orca-mwe.cf.ac.uk/id/eprint/135804

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