Based on density functional theory (DFT), we systematically investigate the formation energies of intrinsic vacancy defects (V_C, V_Si, and V_Si+C) and oxygen-related defects (O_C, O_Si, O_CV_Si, and O_SiV_C) in 3C-SiC. The results indicate that, among the considered defects, all except O_C possess neutral or negative charge states, thereby making them suitable for detection via positron annihilation spectroscopy (PAS). Furthermore, we compute the electron–positron density distributions and positron annihilation lifetimes for the perfect 3C-SiC supercell and various defective configurations. It is found that the O_Si and O_SiV_C complexes serve as effective positron trapping centers, leading to the formation of positron trapped states and a notable increase in annihilation lifetimes at the corresponding defect sites. In addition, coincidence Doppler broadening (CDB) spectra, along with the S and W parameters, are calculated for both intrinsic and oxygen-doped point defects (O_C, O_Si, O_CV_Si, and O_SiV_C). The analysis reveals that electron screening effects dominate the annihilation characteristics of the O_Si defect, whereas positron localization induced by the vacancy is the predominant contributor in the case of O_SiV_C. This distinction results in clearly different momentum distributions for these two oxygen-related defects across various charge states. Overall, PAS is demonstrated to be a powerful technique for distinguishing intrinsic vacancy-type defects from oxygen-doped complexes in 3C-SiC. When combined with electron–positron density analysis, it enables a comprehensive understanding of electron localization and positron trapping behavior in defect systems with different charge states. These first-principles results provide a solid theoretical foundation for the identification and characterization of defects in oxygen-doped 3C-SiC using positron annihilation spectroscopy.