The proposed four-quadrant Moiré alignment scheme to detect the misalignment between mask and wafer for proximity lithography can achieve the alignment accuracy with nanometer level. When implementing the scheme, however, the distribution of Moiré fringes associated with the mask–wafer gap indeed goes against the alignment, making the gap optimization highly urgent. The optimization model is established, and numerical simulation as well as experimental verification is also provided. Furthermore, an alignment accuracy of ∼3 nm with the illumination wavelength of 632.8 nm is experimentally attained. Simultaneously, the design mechanism of alignment marks for improving the availability of the alignment scheme is discussed.