The effects of oxygen content on the microstructure and creep properties of the FGH96 superalloy were investigated. When oxygen content increased from 135 ppm to 341 ppm, the prior particle boundary (PPB) rose from degree 2 to degree 3, the size of the γ′ phase on PPB enlarged from 1.07 μm to 1.27 μm, and the MC carbide size grew from 77.4 nm to 104.0 nm. Meanwhile, the steady creep rate accelerated from 4.34 × 10−3 h−1 to 1.87 × 10−2 h−1, and the creep rupture life shortened from 176 h to 94 h, the creep rupture mode transferred from intergranular and transgranular mixed fracture to along PPB fracture. During creep, the micro-twin formation and gliding will be restrained by ∑3 boundaries. FGH96 superalloy with higher oxygen content contains less ∑3 boundaries, and its micro-twins cross-slipped instead of single-direction slip in lower oxygen content superalloy. Consequently, samples with a higher oxygen content crept faster and ruptured earlier.