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We report that a deionized water leaching and thermal annealing technique can be effective for preparing atomically flat and singly terminated surfaces of single crystalline SrTiO 3 substrates. After a two-step thermal-annealing and deionized-water leaching procedure, topography measured by atomic force microscopy shows the evolution of substrates from a rough to step-terraced surface structure. Lateral force microscopy confirms that the atomically flat surfaces are singly terminated. Moreover, this technique can be used to remove excessive strontium oxide or hydroxide composites segregated on the SrTiO 3 surface. This acid-etchant-free technique facilitates the preparation of atomically aligned SrTiO 3 substrates, which promotes studies on two-dimensional physics of complex oxide interfaces.