The invention provides a separation method and a separation device for needle seat assemblies of a double-needle-handle medical vein needle, and belongs to the technical field of medical equipment. The method and the device solve the technical problems that the needle seat assemblies are low in assembling speed and low in efficiency in the prior art. The separation method comprises the following steps of a, discharge, b, feeding, c, intermittent output, and d, reversing adjustment. The separation device comprises a base, wherein strip-shaped feeding groove bodies used for supporting the needle seat assemblies are fixed on the base; vibrators capable of allowing the feeding groove bodies to generate vibration are arranged between the feeding groove bodies and the base; the needle seat assemblies on the feeding groove bodies can be conveyed forward by the vibration of the feeding groove bodies; strip-shaped pressing strips are fixed on the base, parallel to the feeding groove bodies, and located right above the feeding groove bodies; and gaps capable of allowing the needle seat assemblies to pass through are formed between the pressing strips and the feeding groove bodies. The method and the device have the advantages of high assembling speed, high efficiency and the like.