A laser scanning microscope is capable of separating the fluorescence signals of different fluorophores in accurate unmixing realized by eliminating positional pixels shifts between different fluorescence images obtained through irradiation of different-wavelength laser lights. The microscope includes a laser light source capable of emitting a wavelength-changeable laser light, a correction amount determination unit that determines a correction amount for correcting an optical axis shift of the laser light, an optical axis adjusting unit that adjusts an optical axis, a scanning unit that performs two-dimensional scanning, an objective lens that focuses the laser scanning light to a specimen and fluorescence emitted from the specimen, a light detector that detects the fluorescence, and a control unit that changes the wavelength of the laser light synchronously with the scanning by the scanning unit while controlling the optical axis adjusting unit on the basis of the correction amount determined by correction amount determination unit.