A Cell Culture Model that Mimics Physiological Tissue Oxygenation Using Oxygen-permeable Membranes

Dissolved oxygen and its availability to cells in culture is an overlooked variable which can have significant consequences on experimental research outcomes, including reproducibility. Oxygen sensing pathways play key roles in cell growth and behavior and pericellular oxygen levels should be controlled when establishing in vitro models. Standard cell culture techniques do not have adequate control over pericellular oxygen levels. Slow diffusion through culture media limits the precision of oxygen delivery to cells, making it difficult to accurately reproduce in vivo-like oxygen conditions. Furthermore, different types of cells consume oxygen at varying rates and this can be affected by the density of growing cells. Here, we describe a novel in vitro system that utilizes hypoxic chambers and oxygen-permeable culture dishes to control pericellular oxygen levels and provide rapid oxygen delivery to adherent cells. This procedure is particularly relevant for protocols studying effects of rapid oxygen changes or intermittent hypoxia on cellular behavior. The system is inexpensive and easily assembled without highly specialized equipment.