Amyloid-β (Aβ) containing plaques and tau-laden neurofibrillary tangles are the defining neuropathological features of Alzheimer’s disease (AD). To better mimic this neuropathology, we generated a novel triple transgenic model of AD (3xTg-AD) harboring three mutant genes: β-amyloid precursor protein (βAPP_Swe), presenilin-1 (PS1_M146V), and tau_P301L. The 3xTg-AD mice progressively develop Aβ and tau pathology, with a temporal- and regional-specific profile that closely mimics their development in the human AD brain. We find that Aβ deposits initiate in the cortex and progress to the hippocampus with aging, whereas tau pathology is first apparent in the hippocampus and then progresses to the cortex. Despite equivalent overexpression of the human βAPP and human tau transgenes, Aβ deposition develops prior to the tangle pathology, consistent with the amyloid cascade hypothesis. As these 3xTg-AD mice phenocopy critical aspects of AD neuropathology, this model will be useful in pre-clinical intervention trials, particularly because the efficacy of anti-AD compounds in mitigating the neurodegenerative effects mediated by both signature lesions can be evaluated.