In this study, the thermal oxidative stability of a kerosene-type Jet A-1 commercial aviation fuel has been investigated using a three-dimensional (3D) excitation/emission matrix fluorescence (EEMF) method. The fuel was thermally stressed in flow test conditions over a range of temperatures in the autoxidative regime. To determine the effect of dissolved oxygen on aviation fuel thermal stability, the measurements were conducted for both air-saturated and fully deoxygenated fuel samples. The increase in the fuel temperature results in a large red shift of the fluorescence signals for the air-saturated fuel; however, fully deoxygenated fuel showed no difference in the fluorescence spectra with respect to neat fuel. The observed increase in the emission wavelengths of the collected spectra may be attributed to the formation of high-molecular-mass compounds within the liquid fuel. These species are formed as a consequence of the chemical reactions activated during the thermal stressing process. The use of 3D fluorescence spectra for aviation fuel analysis is shown to be a fast, suitable, and easily implementable tool for establishing and verifying fuel quality. Also, the level of fuel thermal degradation may be ascertained using this method, which should be of great interest for a better thermal management control of turbine engines.