Fullerene bisadducts have emerged as promising electron-accepting materials because of their ability to increase the open-circuit voltage (V-OC) of polymer solar cells (PSCs) due to their relatively high lowest unoccupied molecular orbital (LUMO) energy levels. It should be noted that the as-prepared fullerene bisadducts are in fact a mixture of isomers. Here, the effects of fullerene bisadduct regioisomers on photovoltaic performance are examined. The trans-2, trans-3, trans-4, and e isomers of dihydronaphthyl-based [60]fullerene bisadduct (NCBA) are isolated and used as acceptors for P3HT-based PSCs. The four NCBA isomers exhibit different absorption spectra, electrochemical properties, and electron mobilities, leading to varying PCE values of 5.8, 6.3, 5.6, and 5.5%, respectively, which are higher than that based on an NCBA mixture (5.3%), suggesting the necessity to use the individual fullerene bisadduct isomer for high-performance PSCs.