The invention relates to a method for preparing three-classification whole blood control substance simulacrum and a blood type detection reagent box made by the method. The method comprises the preparation of red blood cell simulacrum, white blood cell simulacrum and blood platelet simulacrum. The concentration of the prepared red blood cell simulacrum, white blood cell simulacrum and blood platelet simulacrum is adjusted to ensure that cell particle is suspended in cell suspension and is used as monomial substance control substance, thereby forming whole blood control substance. The invention aims to solve the source of various cells and provides a method for preparing three-classification whole blood control substance. The invention adopts fish blood to simulate human white blood cell and blood platelet, and waste red blood cell of a livestock red blood cell plasma-mixing station to simulate human red blood cell, thereby simulating the practical histogram distribution of human bloodsample; moreover, the cell suspension adopting the formula of the invention has longer storage life; in addition, the method can carry out simulation operation when adopting the red blood cells of silver carp, grass carp, common carp and crocodile, etc. during experiment, and can obtain excellent quality-control effect through fixative treatment.