The invention discloses a zebrafish embryo heart injecting system and method based on vision servo three-dimensional rotation, and relates to the field of micro-injection. The zebrafish embryo heart injecting system comprises a stereomicroscope, a CCD camera, a supervisory computer, a triaxial mechanical hand, a rotating manipulator, a micropipet and a pump, wherein the CCD camera is located abovethe stereomicroscope, and is used for collecting images and transmitting the collected images to the supervisory computer for treatment; the supervisory computer is connected with the triaxial mechanical hand and the rotating manipulator, and is used for analyzing the real-time position and the gesture of a zebrafish embryo as well as the point position of the micropipet according to the collected images, and transmitting a control signal to the triaxial mechanical hand and the rotating manipulator to control the direction and the speed of the triaxial mechanical hand and the rotating angle of the rotating manipulator; the triaxial mechanical hand is used for clamping the corresponding micropipet to move the point of the corresponding micropipet to a target position; the rotating manipulator is used for clamping the corresponding micropipet to rotate the zebrafish embryo; and the pump is used for providing sucking force for sucking and holding the zebrafish embryo and thrust for pushing out foreign substances. The system is simple in structure, high in precision and good in stability.