A calibration system for robot tool including a robot arm adopting a first coordinate system, a tool arranged on a flange of the robot arm, and an imaging device adopting a second coordinate system is disclosed, wherein an image sensing area is established by the image device. A calibration method is also disclosed and includes steps of: controlling the robot arm to move for leading a tool working point (TWP) of the tool enters the image sensing area; recording a current gesture of the robot arm as well as a specific coordinate of the TWP currently in the second coordinate system; obtaining a transformation matrix previously established for describing a relationship between the first and the second coordinate systems; and importing the specific coordinate and the current gesture to the transformation matrix for calculating an absolute position of the TWP in the first coordinate system.