Graphene oxide sheets (GOSs) of homogeneous size are highly desirable for applications and fundamental research. Herein, we developed a novel, simple and economic method to prepare GOSs of homogeneous size by the modified Hummers method. The method is based on the different dispersibility and stability of varisized GOSs in a polar solvent. The homogeneity in the size of the GOSs obtained can be improved. We found that the dispersibility and stability of GOSs were strongly correlated with the C–O content of GOSs and the lateral dimensions of GOSs decreased while the degree of oxidation increased, which enabled the separation of varisized GOSs. GOSs were separated into four groups, namely GOSs1 (d > 25 μm), GOSs1 (15 μm < d < 25 μm), GOSs3 (5 μm < d < 15 μm) and GOSs4 (<5 μm). The percentages of the size distributions of GOSs1, GOSs2, GOSs3 and GOSs4 reached up to 83.16%, 83.92%, 90.42% and 92.32%, respectively. Furthermore, we also found that the smaller GOSs possessed stronger Rhodamine-B luminescence quenching abilities than the larger GOSs. The more efficient fluorescence resonance energy transfer was deemed to be the major reason for their distinct quenching abilities.