Disclosed herein are systems and methods for constructing an image of a sample using a plurality of images acquired under multiple illumination conditions. In some cases, a microscope may include an image capture device, an illumination assembly, and a processor configured to acquire a plurality of images of a sample and a fiducial marker under a plurality of different illumination conditions and to reconstruct a high resolution image in response to the plurality of images. The disclosure also provides a method for generating a high resolution image of a sample comprising acquiring a plurality of images of a sample and a fiducial marker under a plurality of different illumination conditions and reconstructing the high resolution image in response to the plurality of images.