Measurements of lumbar spine (L1–L6) bone mineral density (BMD) and bone mineral content (BMC) of Wistar rats were obtained by dual energy X-ray absorptiometry (DXA) (QDR-1000W, Hologic Inc., Waltham, MA) to estimate reproducibility and investigate age-related changes. In addition we evaluated the accuracy of the technique in female rats. The coefficients of variation (CV) for spine BMD measurements were found to range from 0.73–1.04 in vivo and from 0.36–1.56 in vitro. The in vitro measurements were performed in a 3 cm deep water bath to stimulate an equivalent tissue thickness. Spine BMC, measured in vivo and in vitro correlated closely with the subsequently determined ash weights (r2 = 0.87 and 0.97, respectively). We examined age-related spine BMD by DXA. A relatively constant increase in spine BMD was observed from 6 weeks to 22 weeks; spine BMD remained stable between 22 and 58 weeks. No peak was observed in spine BMD. To evaluate the effect of estrogen deficiency on animals of different ages, we measured spine BMD weekly in female rats subjected to ovariectomy (OVX) or sham operation at 8 and 23 weeks of age. The spine BMDs in each OVX rat were significantly lower than that of the controls. In the 23-week-old rats, bone loss was quite rapid for the first 3 weeks of observation and stable afterward. The BMD of 8-week-old OVX rats increased with body size. We conclude that DXA allows the observation of age-related changes in the spine BMD of rats with great precision. The data we have gathered using this method should prove useful for the assessment of experimental models of osteoporosis in rats.