Measurements of the speed-of-sound (SOS) and of the broadband ultrasound attenuation (BUA) on the os calcis were recently proposed to assess osteoporotic fragility. Velocity and attenuation were measured through the heel which can be divided in three phases including hydroxyapatite, soft tissue, and fat. The aim of this study was to evaluate the influence of fat composition and heel width on SOS and BUA. This influence was determined from both in vitro investigations examining fat samples, phantoms, and cadaver heels, and in vivo ones observing adult volunteers as well as a wide sample section of healthy elderly women. Ultrasound velocities on various fat samples were significantly lower than those on distilled water (-65 m/second to -123 m/second). The excision of the surrounding soft tissue from cadaver heels made SOS steadily increase whereas the insertion of a 10 mm piece of lard in the lateral face of cadavers' and volunteers' heels os calcis lowered SOS about 30 m/second. Furthermore, a difference of SOS was estimated at 15 m/second for a 12.5% variation of the marrow fat weight. Among 334 elderly and healthy women aged 75 and over, a significant negative correlation was found between SOS and heel width (r = -0.27; P < 0.0001). On the other hand, fat composition had no significant effect on BUA measurement, and no significant relationship was found between BUA and heel width. This study demonstrates that an increase of heel width and fat thickness provides an underestimation of os calcis SOS, but has no significant effect on BUA.