Emissions from agricultural sources substantially contribute to global warming. The inverse dispersion method (IDM) has been successfully used for emission measurements from various agricultural sources. The IDM has also been validated in multiple studies with artificial gas releases mostly in open fields. Release experiments from buildings have rarely been conducted and were partly affected by additional nearby sources of the target gas. Specific release studies for naturally ventilated animal housings are lacking. In this study, a known and predefined amount of methane (CH4) was released from an artificial source inside a barn that mimicked a naturally ventilated dairy housing, and IDM recovery rates, using a backward Lagrangian stochastic (bLS) model, were determined. For concentration measurements, open-path devices (OPs) with a path length of 110m were placed in a downwind direction of the barn at fetches of 2.0h, 5.3h, 8.6h, and 12h (h equals the height of the highest obstacle), and a 3D ultrasonic anemometer (UA) was placed in the middle of the first three OP paths. Upwind of the barn, an additional OP and a UA were installed. The median IDM recovery rates determined with the UA placed upwind of the barn and the downwind OP ranged between 0.55–0.75. It is concluded that, for the present study case, the effect of the building and a tree in the main wind axis led to a systematic underestimation of the IDM-derived emission rate probably due to deviations in the wind field and turbulent dispersion from the underlying assumptions of the used dispersion model.