The present invention discloses a 17beta-estradiol visualization detection method based on DNA nano-structure, and a 17beta-estradiol visualization detection kit based on DNA nano-structure. The working principle is that 17b-estradiol interacts with aptamer, a strand displacement reaction is started, three groups of stem-loop DNA probes are constantly opened to form DNA nano-structures, a G tetramer having catalysis activity is formed on the terminals of the three DNA arms in the DNA nano-structures, the G tetramer is bound with hemin so as to form a compound having catalysis activity and similar to HRP, TMB is subjected to catalytic oxidation, and a blue substrate is produced, wherein the result is visible, and the concentration of 17b-estradiol is directly related to the blue shade. According to the present invention, the operation is simple, the whole reaction can be completed at the room temperature, the high sensitivity is provided, the detection limit on 17b-estradiol is 100 fM, the good specificity is provided, the detection result is directly visible without any detection equipment, and the method and the kit can be used for the on-site rapid detection of 17b-estradiol.