In this study, effects of altitude on free diesel spray morphology, macroscopic spray characteristics and air-fuel mixing process were investigated. The diesel spray visualization experiment using high-speed photography was performed in a constant volume chamber which reproduced the injection diesel-like thermodynamic conditions of a heavy-duty turbocharged diesel engine operating at sea level and 1000 m, 2000 m, 3000 m and 4500 m above sea level. The results showed that the spray morphology became narrower and longer at higher altitude, and small vortex-like structures were observed on the downstream spray periphery. Spray penetration increased and spray angle decreased with increasing altitude. At altitudes of 0 m, 1000 m, 2000 m, 3000 m and 4500 m, the spray penetration at 1.45 ms after start of injection (ASOI) were 79.54 mm, 80.51 mm, 81.49 mm, 83.29 mm and 88.92 mm respectively, and the spray angle were 10.9°, 10.8°, 10.7°, 10.4°and 9.8° respectively. The air-fuel mixing process was analysed using empirical relations like fuel spray area, spray volume, mass of air entrained within the spray and equivalence ratio. The decrease of spray area, spray volume and mass of air entrained were accelerate when altitude increased. At 1000 m, 2000 m, 3000 m and 4500 m, compared with 0 m, the relative changes of spray area were −0.78%, −2.23%, −4.49% and −9.45% respectively, the relative changes of spray volume were −1.54%, −4.38%, −8.72% and −17.9% respectively, and the relative changes of mass of air entrained were −5.06%, −13.18%, −23.89% and −42.68% respectively, showing the negative effect of altitude was more obvious on the mass of air entrained due to the additive effect of spray volume and ambient density. Hence at a given incharge condition of a diesel engine at high altitude, it is necessary to intensify air-fuel mixing to improve oxygen utilization. When altitude increased, the higher axial equivalence ratio showed the quality of air-fuel mixing was deteriorated and similarly the relative changes of axial equivalence ratio also indicated the acceleration of the level of deterioration as altitude rose. Due to the narrower spray angle and smaller mass of air entrained, higher altitude provided larger and sharper distribution of radial equivalence ratio, indicating poor air-fuel mixing at high altitude.