In-situ microscopic studies at air-water interface in terms of optical anisotropy were performed on the two-dimensional crystallines of a polydiacetylene derivative monolayer. Fabrication and evaluation procedures enable observing the structures of various surface crystals, such as spherulites and two-dimensional crystals. Band-shaped crystals were obtained by expansion of surface area under the precision control of the Langmuir trough. We developed for the first time a fabrication method to obtain large two-dimensional single crystals of ca. 20×5mm at the air-water interface.