Retinal surgery is known to be a complicated and challenging task for an ophthalmologist even for retina specialists. Image guided robot-assisted intervention is among the novel and promising solutions that may enhance human capabilities during microsurgery. In this letter, a novel method is proposed for 3D navigation of a microsurgical instrument based on the projection of a spotlight during robot-assisted retinal surgery. To test the feasibility and effectiveness of the proposed method, a vessel tracking task in a phantom with a Remote Center of Motion (RCM) constraint is performed by the Steady-Hand Eye Robot (SHER). The results are compared to manual tracking, cooperative control tracking with the SHER and spotlight-based automatic tracking with SHER. The reported results are that the spotlight-based automatic tracking with SHER can reach an average tracking error of 0.013 mm and keeping distance error of 0.1 mm from the desired range demonstrating a significant improvement compared to manual or cooperative control methods alone.