The invention discloses a tilapia growth hormone quantitative determination kit based on timed-resolved fluoroimmunoassay and heterologous antibody. The kit comprises a micropore reaction plate coated with a saddletail grouper growth hormone specific single chain monoclonal antibody, a recombination tilapia growth hormone protein standard substrate, and an europium-labelled saddletail grouper growth hormone hormone specific single chain monoclonal antibody. Through immunization reaction with a antibody sandwich method, the recombination tilapia growth hormone protein standard substrate or a to-be-detected sample is added in the micropore reaction plate coated with the saddletail grouper growth hormone specific single chain monoclonal antibody, incubation combination is carried out, then board-washing is carried out, then the europium-labelled saddletail grouper growth hormone hormone specific single chain monoclonal antibody is added, incubation combination is carried out, then board-washing is carried out, enhancement liquid is added, and determination is carried out through a multifunctional ELIASA. The kit can be used for detecting growth hormone concentration of growth hormone-containing samples such as tilapia serum or a pituitary lysate, the detection sensitivity can reach 0.225 ng/ml, and the kit provides effective detection means for researching the tilapia growth hormone.