Near-infrared (NIR) quantum dots (QDs) have been treated as a promising candidate of imaging agents for NIR fluorescence-guided surgery. Herein, one-pot synthesis of water-soluble NIR fluorescence CuInS₂ QDs was developed using ribonuclease A (RNase A) as the template. Our product exhibits good colloidal stability, excellent NIR fluorescence properties, and good photostability, even under acidic conditions. Furthermore, the RNase A-CuInS₂ QDs have minor suppression on cell viability even at a concentration of up to 5 mg mL⁻¹. Finally, the following in vivo imaging results confirm that RNase A-CuInS₂ QDs can be used as a novel oral imaging agent in the future.