Based on the first report of the principle and practical observation of additive-type fluorescence moiré fringes for imprint alignment, we here studied the limit accuracy of the fluorescence imprint alignment using image drawing software, image processing software, and data analysis software. According to the practical conditions, fluorescence images of two assemblages having concave lines with a short pitch of p 1 = 4.0 μm and a long pitch of p 2 = 4.4 μm were reproduced on the mold and substrate. The analyzes of reproduced fluorescence images of moving the substrate without Fourier transform suggested that the minimum substrate shift of 0.04 pixels (5.2 nm) could be detected to be 0.045 58 pixels (5.9 nm). The detected placement error had a linear correlation to the substrate shift within 1290 nm.