The invention discloses a guide shaft positioning device which comprises a machine frame, a first guide shaft and a second guide shaft. The guide shaft positioning device further comprises a positioning claw, a first guide base and a second guide base. The first guide base is installed on the first guide shaft in a sliding mode, the second guide base is installed on the second guide shaft in a sliding mode, the two ends of the positioning claw are fixed to the first guide base and the second guide base correspondingly, the positioning claw can slide in the first direction so as to enable the first guide shaft and the second guide shaft to be parallel to each other in the second direction perpendicular to the first direction, and the first guide shaft and the second guide shaft are fixedly installed on the machine frame. The positioning claw can slide in the first direction so as to enable the first guide shaft and the second guide shaft to be parallel to each other in the second direction perpendicular to the first direction, and then the first guide shaft and the second guide shaft are fixedly installed on the machine frame, so that the depth of parallelism of the first guide shaft and the second guide shaft is ensured. The invention further relates to a guide shaft positioning method.