[Object] To propose an information processing device, an image acquisition system, an information processing method, an image information acquisition method, and a program which enable a position of a surface of a measurement subject to be more simply specified.[Solution] An information processing device according to the present invention includes: a representative luminance value specifying unit configured to, when luminance values constituting a plurality of fluorescence images of a measurement subject captured while a position of the measurement subject in a thickness direction is changed are sequentially rearranged from a highest luminance value on the basis of the fluorescence images for each of the fluorescence images corresponding to respective thickness positions, extract a luminance value ranked at a predetermined position from the highest luminance value and set the extracted luminance value as a representative luminance value of the fluorescence image at the thickness position to be noted and a surface position specifying unit configured to use the representative luminance value for each of the fluorescence images and set the thickness position corresponding to the fluorescence image that gives the maximum representative luminance value as a position corresponding to a surface of the measurement subject.
