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Oxygenation in cell culture: Critical parameters for reproducibility are routinely not reported
Open accessAbstract
Mammalian cell culture is foundational to biomedical research, and the reproducibility of research findings across the sciences is drawing increasing attention. While many components contribute to reproducibility, the reporting of factors that impact oxygen delivery in the general biomedical literature has the potential for both significant impact, and immediate improvement. The relationship between the oxygen consumption rate of cells and the diffusive delivery of oxygen through the overlying medium layer means parameters such as medium depth and cell type can cause significant differences in oxygenation for cultures nominally maintained under the same conditions. While oxygenation levels are widely understood to significantly impact the phenotype of cultured cells in the abstract, in practise the importance of the above parameters does not appear to be well recognized in the non-specialist research community. On analyzing two hundred articles from high-impact journals we find a large majority missing at least one key piece of information necessary to ensure consistency in replication. We propose that explicitly reporting these values should be a requirement for publication.
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![Fig 1 Steady-state oxygen mass transfer in cell culture media. Shown in the centre is a plot of the calculated oxygen concentration (mmol/L) across a media column under culture conditions of 0.33¯cm media depth, atmospheric pressure at sea level, a culture density of 200,000 cells / cm2, and an oxygen consumption rate for CHO cells of 8.60×10⁻¹⁷mol/(cell∙s) [47]. Each condition shown around the perimeter represents the consequences of a change in one variable. At the base of each column the green filling within the circle indicates the fraction of the cells’ metabolic oxygen needs that can be met under each condition.](/paper-figures/0b0589cf709e451eb66945848f8bd442/oxygenation-in-cell-culture-critical-parameters-for-reproducibility-are-figure-1.jpg)
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“recent assessments show that the majority of research papers rely on nominal set points and fail to directly verify and report environmental parameters (Al-Ani et al., 2018; Michl et al., 2019; Klein et al., 2021b).”
From Toward Best Practices for Controlling Mammalian Cell Culture Environments (2022) · cites this paper as Al-Ani et al., 2018
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“The lack of monitoring and reporting of environmental conditions in cell culture-systems is a pervasive, but under-recognized problem (Hunter, 2017; Al-Ani et al., 2018; Michl et al., 2019; Klein et al., 2021a; Klein et al., 2021b).”
From Toward Best Practices for Controlling Mammalian Cell Culture Environments (2022) · cites this paper as Al-Ani et al., 2018
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“Medium changes have been shown to influence oxygen supply and cellular metabolism, as well as synthesis of proteins, DNA, and RNA (Al-Ani et al., 2018; Wright Muelas et al., 2018).”
From In Vitro research Reproducibility: Keeping up High Standards (2019) · cites this paper as Al-Ani et al., 2018
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“In addition, the height/level of medium above cells determines the accessibility to oxygen and should be recognized as an important factor for cell culture (Al-Ani et al., 2018).”
From Guidance document on Good cell and tissue culture practice 2.0 (GCCP 2.0) (2021) · cites this paper as Al-Ani et al., 2018
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“In the case of O2, medium deoxygenation is caused by the disparity between rates of O2 consumption via cellular metabolism and the replenishment of O2 at the air-medium interface (Place et al., 2017; Al-Ani et al., 2018).”
From Toward Best Practices for Controlling Mammalian Cell Culture Environments (2022) · cites this paper as Al-Ani et al., 2018
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“although reliable reports of environmental parameters within selected human tissues are presently limited (Al-Ani et al., 2018; Ast and Mootha, 2019)”
From Toward Best Practices for Controlling Mammalian Cell Culture Environments (2022) · cites this paper as Al-Ani et al., 2018
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Metadata
- Authors
- Al-Ani, A., Toms, D., Kondro, D., Thundathil, J., Yu, Y., & Ungrin, M.
- Affiliations
- University of Alberta (CA)
- University of Calgary (CA)
- Corresponding author
- Al-Ani, Abdullah
- Toms, Derek
- Kondro, Douglas
- Thundathil, Jarin
- Yu, Yang
- Ungrin, Mark
- Venue
- PLOS ONE
- Publisher
- Public Library of Science (PLoS)
- Published
- 2018-10-16
- Volume
- 13
- Issue
- 10
- Pages
- e0204269
- ISSN
- 1932-6203 (ISSN-L)
- PMC
- PMC6191109
- OpenAlex
- W2896573442
- Cited by
- 152
- References
- 76
- Citations by year
- 2026: 15 · 2025: 20 · 2024: 15 · 2023: 25 · 2022: 29 · 2021: 25 · 2020: 15 · 2019: 8
- FWCI
- 3.63
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- Locations
- PLoS ONE (publishedVersion, cc-by)
- Europe PMC (PubMed Central) (publishedVersion, cc-by)
- PubMed Central (publishedVersion, cc-by)
- PubMed (publishedVersion)
- RePEc: Research Papers in Economics (submittedVersion)
- DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion)
- Figshare (submittedVersion, cc-by)
- Funders
- Natural Sciences and Engineering Research Council of Canada (RGPIN-201404874, Graduate Studentship)
- Institute of Nutrition, Metabolism and Diabetes (MOP-137095)
- Alberta Diabetes Institute (Graduate Studentship)
- Alberta Children's Hospital Research Institute (CA) (Graduate Studentship)
- Alberta Children's Hospital Research Institute (Post-doctoral Fellowship)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Cancer Research › Cancer, Hypoxia, and Metabolism
- Health Sciences › Veterinary › Small Animals › Animal testing and alternatives
- Social Sciences › Library and Information Sciences › Academic Writing and Publishing
- Keywords
- oxygenation, mammalian cell culture, oxygen consumption rate, oxygen diffusion, reproducibility, research reporting practices, publication requirements, culture medium composition, cell phenotype
- MeSH terms
- Animals, Cells, Cultured, Humans, Mammals, Oxygen, Phenotype, Research Design, Cell Culture Techniques
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