Oxygenation in cell culture: Critical parameters for reproducibility are routinely not reported

Abstract

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.

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 $0.3\overline{3}\mathrm{c}\mathrm{m}$ media depth, atmospheric pressure at sea level, a culture density of 200,000 cells / cm², and an oxygen consumption rate for CHO cells of $8.60\times {10}^{-17}\frac{\mathrm{m}\mathrm{o}\mathrm{l}}{\mathrm{c}\mathrm{e}\mathrm{l}\mathrm{l}\bullet \mathrm{s}}$ [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.

Fig 2. Results from analysis of published work employing mammalian cell culture.

Manuscripts were assessed and identified as specifying all of the critical variables (blue), missing only one of the critical variables (orange), or missing multiple critical variables (grey).