This paper reports a novel 3D (three dimensional) reconstruction method from optical microscopic images. The method can effectively reconstruct 3D microstructures based on digital image processing and precision mechanics, greater than the field of view of an optical microscope of 0.647 mm × 0.483 mm. Firstly, partial-sample images are taken on different focal planes for the whole microstructure. Those partial-sample images are merged to create a sequence of whole-sample images of different focal planes. The sequence of whole-sample images is spliced to keep only the area of interests by using a highly efficient image splicing methods. The spliced images sequence is then used in 3D reconstruction of the microstructure by a coincidence algorithm. Experimental results show that the method is simple and effective, and the 3D reconstructed image provides richer information of the microstructure.