An experimental and numerical investigation on natural convection heat transfer of TiO2–water nanofluids in a square enclosure was carried out for the present work. TiO2–water nanofluids with different nanoparticle mass fractions were prepared for the experiment and physical properties of the nanofluids including thermal conductivity and viscosity were measured. Results show that both thermal conductivity and viscosity increase when increasing the mass fraction of TiO2 nanoparticles. In addition, the thermal conductivity of nanofluids increases, while the viscosity of nanofluids decreases with increasing the temperature. Nusselt numbers under different Rayleigh numbers were obtained from experimental data. Experimental results show that natural convection heat transfer of nanofluids is no better than water and even worse when the Rayleigh number is low. Numerical studies are carried out by a Lattice Boltzmann model (LBM) coupling the density and the temperature distribution functions to simulate the convection heat transfer in the enclosure. The experimental and numerical results are compared with each other finding a good match in this investigation, and the results indicate that natural convection heat transfer of TiO2–water nanofluids is more sensitive to viscosity than to thermal conductivity.