Bone is mainly composed of hydroxyapatite (principally carbonated apatite) and type I collagen. We have previously demonstrated that collagen fiber orientation in the rat femoral cortical bone contributed its bone strength using IR dichroism imaging in transmittance mode. In this work, we characterized collagen fiber orientation in bone embedded in a PMMA block as well as in thin sections of bone using IR dichroism imaging in reflectance mode in order to assess the same samples by various analytical methods, and demonstrated that the distribution of collagen fiber orientation observed in reflectance mode coincided with that in transmittance mode.