Abstract There is relatively little literature concerning the effect of Reynolds number on multi-hole aerodynamic probe performance. In particular, there is almost no discussion in the literature regarding the underlying mechanisms of Reynolds number (Re) sensitivity for such probes. In order to close this gap, detailed investigations of the effect of Re on a five-hole probe have been performed using both particle image velocimetry (PIV) techniques and oil flow visualizations. Wind- and water-tunnels were used to cover a wide range of Re. The open-access Oxford Probe was used for these studies because of the readily available data-sets and processing routines and to allow future comparisons by other authors. Complex flow dynamics including flow separation and re-attachment were identified, which cause Re-sensitivity of the calibration map at low Re even for low yaw or pitch angles. By comparing calibration maps across a wide range of Re, we demonstrate that the Oxford Probe can be employed without much loss of accuracy at lower Re levels than initially (conservatively) suggested, and quantify the errors in the extreme low-Re regime. Overall, we demonstrate the robustness of the Oxford Probe concept across a wide range of Re conditions, we more clearly defined the low-Re limit for the probe design and quantify errors below this limit, and we illustrate the fundamental mechanisms for Re-sensitivity of multi-hole probes.