Background Cerebral specialization and interhemispheric cooperation are two vital features of the human brain. Their dysfunction may be associated with disease progression in patients with Alzheimer's disease (AD), which is featured as progressive cognitive degeneration and asymmetric neuropathology. Objective This study aimed to examine and define two inherent properties of hemispheric function in patients with AD by utilizing resting‐state functional magnetic resonance imaging (rs‐fMRI). Methods Sixty‐four clinically diagnosed AD patients and 52 age‐ and sex‐matched cognitively normal subjects were recruited and underwent MRI and clinical evaluation. We calculated and compared brain specialization (autonomy index, AI) and interhemispheric cooperation (connectivity between functionally homotopic voxels, CFH). Results In comparison to healthy controls, patients with AD exhibited enhanced AI in the left middle occipital gyrus. This increase in specialization can be attributed to reduced functional connectivity in the contralateral region, such as the right temporal lobe. The CFH of the bilateral precuneus and prefrontal areas was significantly decreased in AD patients compared to controls. Imaging‐cognitive correlation analysis indicated that the CFH of the right prefrontal cortex was marginally positively related to the Montreal Cognitive Assessment score in patients and the Auditory Verbal Learning Test score. Moreover, taking abnormal AI and CFH values as features, support vector machine‐based classification achieved good accuracy, sensitivity, specificity, and area under the curve by leave‐one‐out cross‐validation. Conclusion This study suggests that individuals with AD have abnormal cerebral specialization and interhemispheric cooperation. This provides new insights for further elucidation of the pathological mechanisms of AD. Dysfunction of specialization(AI)and interhemispheric cooperation (CFH) was observed in our patients, which may comprise a potential neural substrate for cognitive impairment during AD progression.