Shortcomings of previously reported preclinical models of non-alcoholic steatohepatitis (NASH) include inadequate methods used to induce disease and assess liver pathology. We have developed a dietary model of NASH displaying features observed clinically, and methods for objectively assessing disease progression. Mice fed a diet containing 40% fat (of which ~18% was trans-fat), 22% fructose, and 2% cholesterol developed three stages of non-alcoholic fatty liver disease (steatosis, steatohepatitis with fibrosis, and cirrhosis) as assessed by histological and biochemical methods. Using digital pathology to reconstruct the left lateral and right medial lobes of the liver, comparisons between and within lobes were made to determine the uniformity of collagen deposition, and in turn informed experimental sampling methods for histological, biochemical, and gene expression analyses. Gene-expression analyses conducted using animals stratified by disease severity led to the identification of several genes for which expression highly correlated with the histological assessment of fibrosis. Importantly, we have established a biopsy method allowing assessment of disease progression. Mice subjected to liver biopsy recovered well from the procedure when compared to sham-operated controls with no apparent effect on liver function. Tissue obtained by biopsy was sufficient for gene and protein expression analyses, providing the opportunity to establish an objective method of assessing liver pathology before subjecting animals to treatment. The improved assessment techniques, and the observation that mice fed the high fat diet exhibit many clinically relevant characteristics of NASH establishes a preclinical model for identifying pharmacological interventions with greater likelihood of translating to the clinic.