In contrast to birds, bats are possibly limited in their capacity to use body fat as an energy source for long migrations. Here, we studied the fuel choice of migratory Pipistrellus nathusii (approximate weight: 8 g) by analysing the stable carbon isotope ratio (δ(13)C(V-PDB)) of breath and potential energy sources. Breath δ(13)C(V-PDB) was intermediate between δ(13)C(V-PDB) of insect prey and adipocyte triacylglycerols, suggesting a mixed-fuel use of P. nathusii during autumn migration. To clarify the origin of oxidized fatty acids, we performed feeding experiments with captive P. nathusii. After an insect diet, bat breath was enriched in (13)C relative to the bulk and fat portion of insects, but not deviating from the non-fat portion of insects, suggesting that bats oxidized exogenous proteins and carbohydrates, but not exogenous fatty acids. A feeding experiment with (13)C-labelled substrates confirmed these findings. In conclusion, migratory P. nathusii oxidized dietary proteins directly from insects captured en route in combination with endogenous fatty acids from adipocytes, and replenished their body reserves by routing dietary fatty acids to their body reserves.