Microalgae have been considered as a promising biosorbent for recovery of precious metal ions from diluted aqueous solutions; especially the self-flocculating microalga has the potential for cost-effective harvesting of tiny microalgae. In this study, the adsorption of Au(III) by the self-flocculating microalgae Tetradesmus obliquus and non-flocculating T. obliquus as a control was studied in different initial Au(III) concentrations, temperatures, and pH. The adsorption equilibrium by the tested microalgae fitted well to the Langmuir model at different temperatures. The maximum adsorption capacity from the Langmuir model for T. obliquus AS-6-1 was 169.49 mg g−1 obtained at pH 2.0 and 30 °C, which was higher than that observed by the non-flocculating T. obliquus with 153.85 mg g−1. Furthermore, all the Kd values above 5000 mL g−1 at 30 °C for T. obliquus AS-6-1 showed that the flocculating microalgae had a stronger adsorption affinity to Au(III). The initial Au(III) concentration and solution pH significantly affected the adsorption capacity of Au(III) on algal species. While, their adsorption of Au(III) was slightly influenced at temperature from 30 to 40 °C but significantly influenced and decreased at low (≤ 7 °C) or high temperature (≥ 60 °C). The experimental data fitted well to the second-order kinetic model, which indicated that the adsorption reaction on the surface of adsorbent was the rate-limiting step, instead of mass transfer.