Confocal laser endomicroscopy technology can obtain cell-level images in real time and in situ, which can assist doctors in real-time intraoperative diagnosis, but its non-invasiveness makes it difficult to relocate the optical biopsy site. The confocal probe localization algorithm can automatically calculate the coordinates of the probe tip, that is, the coordinates of the optical biopsy site. In this paper, a confocal probe localization algorithm based on region growing and endoscope size prior was proposed. The algorithm detected the probe region by region growing on the probe edge image, then searched for tip points based on a given probe axis, and iteratively optimized it. Finally, based on the single-degree-of-freedom motion characteristics of the probe, the three-dimensional coordinates of the tip of the probe were calculated by using the prior information of the size of the endoscope, which solved the scale uncertainty problem of the monocular camera. The confocal probe localization algorithm was tested on the dataset collected in this paper. The results showed that our algorithm no longer relied on the color information of the probe, avoided the influence of uneven illumination on the gray value of the probe pixels, and had a more robust location accuracy and running speed. Within the length of the probe extending out of the endoscope from 0 to 5 cm, the pixel error could be as low as 11.76 pixels, and the average relative position error could be as low as 1.66 mm, which can achieve the real-time and accurate localization of the confocal probe.