In the midst of an obesity epidemic in humans, diet induced obesity studies in rodents are fundamental to unravel the complex mechanisms underlying this disease, ultimately resulting in the identification of new preventative and therapeutic strategies. The current study was designed to determine if high throughput multiobject CT scanning was capable of providing precise quantification of adipose tissue in C57BL/6 mice when benchmarked to the gold standard method for evaluating fat mass (freeze drying). We report a strong correlation between body weight alone and fat percentage in our mouse cohort (20 g-40 g, r = 0.95). The gonadal fat depot was identified as the most accurate single predictor of total fat mass (r = 0.931). Importantly, we observed a high positive correlation between both live tissue weight and dissected adipose tissue when correlated to CT predictions (r ≥ 0.862), suggesting CT can accurately be used to predict total fat mass/percentage and non-fat mass/percentage in our cohort. We conclude that the use of multi-object in vivo CT fat quantification is cost effective, accurate and minimally invasive technique in the genetic manipulation era to exploit lean/obese genes in the study of diet induced obesity, allowing longitudinal studies to be completed in a high throughput manner.