Tumor blood flow (TBF) is related to drug delivery and hypoxia, both of which can impact the efficacy of anti-cancer therapies. Although integrin α_vβ₃ expression is related to tumor angiogenesis, it remains unclear whether the degree of angiogenesis affects TBF. This study aimed to evaluate the expression of integrin α_vβ₃ in mouse tumor models using [⁶⁸Ga]Ga-DOTA-c(RGDfK) peptide positron emission tomography (PET) and immunohistochemical staining. PET studies were conducted using mouse C6 glioma models and MIA PaCa-2 (n = 6 each). The [⁶⁸Ga]Ga-DOTA-c(RGDfK) peptide was injected via the tail vein (2.17 ± 0.28 MBq), and 10 min static PET scans were performed. Immunohistochemical analysis was conducted using an integrin α_Vβ₃ antibody. [⁶⁸Ga]Ga-DOTA-c(RGDfK) peptide PET revealed higher uptake of the radiotracer in C6 gliomas than in MIA PaCa-2 tumors. The mean standardized uptake value was significantly higher in C6 gliomas (0.35 ± 0.058) than in MIA PaCa-2 tumors (0.17 ± 0.045). Histological analysis revealed intense integrin α_Vβ₃ expression in the C6 gliomas, whereas the MIA PaCa-2 tumors had low expression levels. This study showed that the expression of integrin α_vβ₃ can be differentiated by the [⁶⁸Ga]Ga-DOTA-c(RGDfK) peptide, suggesting the potential applicability of this peptide in the evaluation of the relationship between angiogenesis and TBF.