This fluorescence observation method is a method of observing a living organism into which a fluorescent dye is injected. The method includes the steps of: irradiating the living organism with excitation light including a wavelength for exciting the fluorescent dye using a light irradiation means, acquiring a first fluorescence image of the living organism generated by the irradiation with the excitation light using an image acquisition means, specifying an observation object in the living organism on the basis of the first fluorescence image； acquiring a second fluorescence image of the observation object generated by the irradiation with the excitation light using the image acquisition means； and specifying a linear fluorescence pattern appearing in the second fluorescence image.
