The spray and combustion of diesel fuel were investigated to provide a better understanding of the evaporation and combustion process under the simulated cold-start condition of a diesel engine. The experiment was conducted in a constant volume combustion chamber and the engine cranking period was selected as the target ambient condition. Mie scattering and shadowgraph techniques were used to visualize the liquid- and vapor-phase of the fuel under evaporating non-combustion conditions (oxygen concentration=0%). In-chamber pressure and direct flame visualization were acquired for spray combustion conditions (oxygen concentration=21%). The fuel was injected at an injection pressure of 30 MPa, which is the typical pressure during the cranking period. The liquid length of the fuel at an ambient temperature of 573 K increased by about 14% compared to that at 663 K due to the lower ambient temperature and fuel temperature as well as the increased fuel density and viscosity from the lower fuel temperature. The vapor penetration of the fuel was also slightly increased at 573 K. However, the vapor phase area was reduced by about 30% at 573 K compared to that at 663 K, which is due to the deteriorated spray atomization and the lower ambient temperature. In the spray combustion condition, only part of the diesel spray was ignited at 573 K, while all of the spray was successfully ignited at 663 K. The ignition delay was increased at 573 K due to the increased injection delay, the deteriorated atomization, and the evaporation of the fuel. The heat release from the premixed combustion was reduced at 573 K, despite the increased ignition delay. This can be explained by the limited formation of the combustible fuel-air mixture as indicated by a reduced vapor-phase area. The heat release from the mixing-controlled combustion was also reduced at 573 K, which resulted in lowering the in-chamber pressure rise by about 42% compared to 663 K.