Thermo-responsiveness and self-assembly behaviors of poly(sodium-p-styrene sulfonate)-block-poly (N-isopropylacrylamide) (PSSS50-b-PNIPAM300) in pure water and binary solvents (i.e., water/methanol and water/acetone) were investigated by 1H NMR, dynamic light scattering (DLS), and transmission electron microscopy (TEM). It was found that the PNIPAM chains had a lower lowest critical solution temperature (LCST) in the water/methanol and water/acetone binary solvents than in water. Temperature-induced collapse of PNIPAM in water/acetone binary solvent was evidently weaker compared to the collapses in pure water and water/methanol binary solvent. The DLS and TEM results demonstrated that the self-assembly process and morphology of PSSS50-b-PNIPAM300 in aqueous solution could be fine-tuned with the addition of small molecule solvent.