A region in which many fluorescent substances are present is identified without carrying out a complicated calculation even if a fluorescence intensity has changed due to a distance and an angle. Provided is a fluorescence observation device (1) comprising an illuminating section (3) that emits excitation light and illumination light onto a subject (A) a fluorescence-image capturing section (18) that acquires a fluorescence image (S2) by image-capturing fluorescence generated in the subject (A) by irradiation with the excitation light from the illuminating section (3) a return-light-image capturing section (17) that acquires a return-light image (S1) by image-capturing return light that returns from the subject (A) by irradiation with the illumination light from the illuminating section (3) a region extracting section (24) that extracts a high-luminance region in which the fluorescence image (S2) acquired by the fluorescence-image capturing section (18) has a gradation level higher than or equal to a set threshold value (S4) and a threshold setting section (22) that sets the threshold value (S4) higher as the gradation level of the return-light image (S1) acquired by the return-light-image capturing section (17) increases.
