A fluorescence image registration method includes obtaining at least one fluorescence image of a biochip. An interior local area. Sums of pixel values in the interior local area along a first direction and a second direction are obtained. A plurality of first template lines is selected to find a minimum total value of the sums of pixel values corresponding to the first template lines. Pixel-level correction is performed on a local area of the track line to obtain a pixel-level track cross. Other track crosses on the biochip is obtained, and the pixel-level correction is performed on the other track crosses. The position of the pixel-level track line is corrected by a center-of-gravity method to obtain the subpixel-level position of the track line. The subpixel-level positions of all sites uniformly distributed on the biochip is obtained.