In the present study, the proportion of nanofluid and dispersion of CuO nanoparticles in ethylene glycol (EG)–water mixture are considered. The effects of pH (in the constant temperature), weight percent of the nanoparticles, and the temperature are also studied. The zeta potential, absorbance and thermal conductivity are measured using a Malvern ZS Nano S analyzer, the UV–visible Spectroscopy, and the Transient Hot Wire (THW) methods. Moreover, in order to investigate the dispersion behavior, the heat transfer enhancement of the nanofluid and the thermal properties of the CuO nanoparticles are measured. Of course, the rate of the obtained thermal conductivity is in the 20–95°C range. The results show that CuO nanoparticles have a significant thermal effect on the mixture. In the optimum pH 7.8, the nanofluid has a maximum zeta potential and a maximum absorbance that can lead to a more stable state. Furthermore, in this optimum pH, the percent of optimum weight and the thermal conductivity enhancement (at 25°C) are 0.045 and 1.55, respectively. Also, the optimum thermal conductivity enhancement at 70°C is 1.66.