Polymer nanospheres are commonly used in scientific and industrial fields including petroleum, electronics, and biomedicine. The precise synthesis of polymer nanospheres with a controllable size and regular shape is important for achieving thermal stability and mechanical properties, which will directly affect the oil displacement effect. In this paper, amphoteric CP(AM-AMPS-DMDAAC) nanospheres with a small particle size and regular shape were prepared by using an acrylamide (AM), 2-acrylamide-2methylpropanesulfonic acid (AMPS), diallyl dimethylammonium chloride (DMDAAC), and methylene-bis-acrylamide mixed solution as the aqueous phase and Span-80 and Tween-80 as the composite emulsifier, followed by reverse-phase microemulsion polymerization. The results showed that nanospheres had temperature and salt resistance and excellent physical and chemical properties. The thermal decomposition temperature was 236.5 °C, the median size of the particles was close to 50 nm, the swelling factor remained at 6.1 nm/nm after water absorption and expansion in the formation water at 120 °C for 7 days, and the swelling factor was 10.4 nm/nm after 14 days. In addition, nanospheres also exhibit good viscoelasticity and pseudoplasticity. The average plugging rate was 79.5% in the low permeability cores, and after the injection of the CP(AM-AMPS-DMDAAC) nanospheres based on water flooding, the average enhanced recovery rate was 13.46%.