Two-Dimensional Structured Illumination System for Epi-Illumination Microscopy

In this work, a two-dimensional structured illumination system integrated into an epi-illumination microscope was designed, constructed, and calibrated. A DMD projector was used as the illumination source, together with a lens system consisting of a camera lens and an achromatic biconvex lens, designed to collimate the white-light beam entering the microscope’s rear aperture. A Python-based algorithm was developed to acquire sample images, segment them using the Segment Anything Model (SAM), and generate masks based on color similarity using the K-means clustering algorithm. To ensure spatial alignment between the projected mask and the sample, the system was calibrated using a homography transformation. For both the 4× and 10× microscope objectives, wavelength-selective illumination was successfully projected onto a square sample region. However, significant chromatic aberration was observed in the projection, which represents an aspect to be addressed in future work.