Evidence-based guidelines for controlling pH in mammalian live-cell culture systems

Abstract

A fundamental variable in culture medium is its pH, which must be controlled by an appropriately formulated buffering regime, since biological processes are exquisitely sensitive to acid–base chemistry. Although awareness of the importance of pH is fostered early in the training of researchers, there are no consensus guidelines for best practice in managing pH in cell cultures, and reporting standards relating to pH are typically inadequate. Furthermore, many laboratories adopt bespoke approaches to controlling pH, some of which inadvertently produce artefacts that increase noise, compromise reproducibility or lead to the misinterpretation of data. Here, we use real-time measurements of medium pH and intracellular pH under live-cell culture conditions to describe the effects of various buffering regimes, including physiological CO ₂/HCO ₃ ⁻ and non-volatile buffers (e.g. HEPES). We highlight those cases that result in poor control, non-intuitive outcomes and erroneous inferences. To improve data reproducibility, we propose guidelines for controlling pH in culture systems.

Abstract

Johanna Michl et al. use real-time pH measurements of cell culture media and intracellular pH data from live-cell culture conditions to explore the effects of commonly-used buffers. Based on these data, they propose guidelines for controlling pH in cell culture systems and improving reproducibility.