A fabrication method of a packaging substrate is provided, which includes the steps of: forming first conductive portions on a carrier; sequentially forming a conductive post and an alignment layer on each of the first conductive portions; forming an encapsulant on the carrier for encapsulating the first conductive portions, the conductive posts and the alignment layers; forming a conductive via on each of the alignment layers in the encapsulant and forming second conductive portions on the conductive vias and the encapsulant; and removing the carrier. Each of the first conductive portions and the corresponding conductive post, the alignment layer and the conductive via form a conductive structure. The alignment layer has a vertical projection area larger than those of the conductive post and the conductive via to thereby reduce the size of the conductive post and the conductive via, thus increasing the wiring density and the electronic element mounting density.