A manufacturing method of multilayer printed circuit boards has steps as follows: aligning circuit layers with a stacking location to form a substrate having multiple positioning portions; pre joining the substrate at the positioning portions; forming an alignment hole in each positioning portion; and placing the pre joined substrate over alignment pins of a press device for lamination. After the circuit layers are aligned, the substrate is pre joined at the positioning portions, and then the alignment holes are formed in the positioning portions for pins alignment at the press device. The alignment accuracy is enhanced. Dusts will not deposit onto surfaces of the circuit layers to damage circuits thereof.