The present invention belongs to the technical field of analysis and detection and provides a method for detecting a selenoamino acid in a selenoprotein. The detection method includes: (1): mixing 10-30 mg of a selenoprotein sample with hydrochloric acid, and hydrolyzing with microwaves at 140-170° C. for 10-40 min to obtain a hydrolysate; (2): adjusting pH of the hydrolysate to 6-8 to obtain a pretreated solution; and, (3): detecting a selenoamino acid in the pretreated solution with high performance liquid chromatography-hydride generation-atomic fluorescence spectrometry (HPLC-HG-AFS) to obtain a content of the selenoamino acid. The method is simple in operation and provides an accurate and reliable result, and also reduces time and cost.