The mixing height (MH) is a crucial parameter in commonly used transport models that proportionally affects air concentrations of trace gases with sources/sinks near the ground and on diurnal scales. Past synthetic data experiments indicated the possibility to improve tracer transport by minimizing errors of simulated MHs. In this paper we evaluate a method to constrain the Lagrangian particle dispersion model STILT (Stochastic Time-Inverted Lagrangian Transport) with MH diagnosed from radiosonde profiles using a bulk Richardson method. The same method was used to obtain hourly MHs for the period September/October 2009 from the Weather Research and Forecasting (WRF) model, which covers the European continent at 10 km horizontal resolution. Kriging with external drift (KED) was applied to estimate optimized MHs from observed and modelled MHs, which were used as input for STILT to assess the impact on CO₂ transport. Special care has been taken to account for uncertainty in MH retrieval in this estimation process. MHs and CO₂ concentrations were compared to vertical profiles from aircraft in situ data. We put an emphasis on testing the consistency of estimated MHs to observed vertical mixing of CO₂. Modelled CO₂ was also compared with continuous measurements made at Cabauw and Heidelberg stations. WRF MHs were significantly biased by ~10–20% during day and ~40–60% during night. Optimized MHs reduced this bias to ~5% with additional slight improvements in random errors. The KED MHs were generally more consistent with observed CO₂ mixing. The use of optimized MHs had in general a favourable impact on CO₂ transport, with bias reductions of 5–45% (day) and 60–90% (night). This indicates that a large part of the found CO₂ model–data mismatch was indeed due to MH errors. Other causes for CO₂ mismatch are discussed. Applicability of our method is discussed in the context of CO₂ inversions at regional scales.