Precise control of dissolved oxygen is essential for reproducing physiologically relevant conditions in microfluidic cell culture systems. Here, we present a standalone, polydimethylsiloxane-free gas exchange unit (GEU) which enables controlled oxygenation and deoxygenation of perfused liquids and is suitable for integration with existing microfluidic platforms. The GEU employs a flat-sheet membrane contactor design to achieve efficient gas–liquid mass transfer while remaining independent of the downstream device. Oxygen transfer was experimentally characterised using optical oxygen sensors under different liquid and gas flow conditions. Reoxygenation efficiency decreased with increasing liquid flow rate because of reduced residence time, whereas active airflow through the gas compartment enhanced oxygen transfer. Controlled deoxygenation was achieved by flowing nitrogen through the gas compartment, with higher nitrogen pressures producing progressively lower oxygen levels. Numerical flow simulations demonstrated uniform flow distribution within the device, and a simplified analytical diffusion model accurately predicted the observed oxygen transfer trends. The proposed GEU provides a simple, robust and modular strategy for regulating dissolved oxygen upstream of microfluidic devices without requiring device redesign or specialised fabrication. This approach offers a practical solution for incorporating physiologically relevant oxygen control into a wide range of microfluidic and cell culture applications.