Influences of base metal and welding speed on fusion zone microstructure and heat affected zone (HAZ) hot-cracking of electron beam welded (EBW) Inconel 718 were investigated systematically. Microstructures of fusion zone and HAZ were characterized using the optical microscope and a field emission scanning electron microscope, complemented by the microhardness measurements. It is found that the weld shape transforms from a stemless wine glass shape to a nail head shape with the increase of the grain size of base metal. The largest amount of Laves or the lowest hardness is obtained in the center of the weld pool, which mainly due to the strong microsegregation in this region. The microsegregation of weld pool is significantly affected by welding speed, which is associated with the rate of transference of energy to the weld. The occurrence of HAZ cracking is related to the grain boundary and the generation of porosities is induced by fracture of the brittle carbides. Furthermore, the susceptibility of HAZ cracking is greatly dependent on the grain size of base metal and welding speed.