Objective: To determine whether oxygen (O2) tension (20% vs. 5%) has an impact on oocyte maturation rates and morphology during in vitro maturation (IVM). Design: A prospective, observational, monocentric, sibling-oocyte study. Setting: University Hospital. Patients: A total of 143 patients who underwent IVM for fertility preservation purposes from November 2016 to April 2021 were analyzed. Patients were included when ≥2 cumulus-oocyte complexes (COCs) were retrieved. The cohort of COCs obtained for each patient was randomly split into two groups: group 20% O2 and group 5% O2. Intervention: Cumulus-oocyte complexes were incubated for 48 hours either under 5% O2 or 20% O2. After 24 and 48 hours of culture, every oocyte was assessed for maturity and morphology, to estimate oocyte quality. Morphology was evaluated considering six parameters (shape, size, ooplasm, perivitelline space, zona pellucida, and polar body characteristics), giving a total oocyte score ranging from -6 to +6. Main outcome measures: Maturation rates and total oocyte scores were compared using paired-sample analysis between group 20% O2 and group 5% O2. Results: Patient median age was 31.4 [28.1-35.2] years-old. The mean serum antimüllerian hormone levels and antral follicle count were 3.2 ± 2.3 ng/mL and 27.2 ± 16.0 follicles, respectively. A mean of 10.7 COCs per cycle were retrieved, leading to 6.1 ± 2.4 metaphase II oocytes vitrified (total maturation rate = 57.3%; 991 metaphase II oocytes/1,728 COCs). A total of 864 COCs were included in each group. Oocyte maturation rates were not different between the two groups (group 20% O2: 56.82% vs. group 5% O2: 57.87%, respectively). Regarding oocyte morphology, the mean total oocyte score was significantly higher in group 5% O2 compared with group 20% O2 (3.44 ± 1.26 vs. 3.16 ± 1.32, P=.014). Conclusion: As culture under low O2 tension (5% O2) improves oocyte morphology IVM, our results suggest that culture under hypoxia should be standardized. Additional studies are warranted to assess the impact of O2 tension on oocyte maturation and the benefit of IVM under low O2 tension for embryo culture after utilization of frozen material.