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The dentate gyrus (DG) is an integral portion of the hippocampal formation, and it is composed of three layers. Quantitative magnetic resonance (MR) imaging has the capability to map brain tissue microstructural properties which can be exploited to investigate neurodegeneration in Alzheimer’s disease (AD). However, assessing subtle pathological changes within layers requires high resolution imaging and histological validation. In this study, we utilized a 16.4 Tesla scanner to acquire ex vivo multi-parameter quantitative MRI measures in human specimens across the layers of the DG. Using quantitative diffusion tensor imaging (DTI) and multi-parameter MR measurements acquired from AD (N = 4) and cognitively normal control (N = 6) tissues, we performed correlation analyses with histological measurements. Here, we found that quantitative MRI measures were significantly correlated with neurofilament and phosphorylated Tau density, suggesting sensitivity to layer-specific changes in the DG of AD tissues.