Micro-Raman spectroscopy (MRS) is used for the first time to our knowledge to investigate brain hippocampus tissue from Alzheimer’s disease (AD) infected rats. In situ Raman analysis of tissue sections provides distinct spectra useful for distinguishing AD from normal state. The biochemical changes of brain hippocampus tissue including the deposit of β-amyloid ( $A\beta$ ) protein, the increase of cholesterol, and hyperphosphorylated tau are observed through MRS when AD occurs. A more convincing multi- Raman criterion based on single Raman peaks, and further in combination with statistical analysis of the entire Raman spectrum, is found capable of classifying brain hippocampus tissues with different pathological features. This study demonstrates the brain hippocampus is an important candidate for considering the early pathological state of AD, and Raman signatures from the brain hippocampus could aid AD diagnosis. In addition, Raman results undoubtedly confirm simultaneous changes of cholesterol and $A\beta$ in the progression of AD.