The present invention discloses an enzyme-linked immunosorbent assay kit for detecting dinitolmide and use thereof. The enzyme-linked immunosorbent assay kit of the present invention comprises dinitolmide antibody and coating antigen and enzyme conjugate; wherein, the dinitolmide antibody is dinitolmide monoclonal antibody or dinitolmide polyclonal antibody; when the coating antigen is the conjugate of dinitolmide hapten and carrier protein, the enzyme conjugate is enzyme-labeled secondary antibody, or enzyme-labeled specific anti-dinitolmide monoclonal or polyclonal antibody; when the coating antigen is dinitolmide antibody or secondary antibody, the enzyme conjugate is enzyme-labeled dinitolmide hapten. The enzyme-linked immunosorbent assay kit has a simple structure, and it is convenient for use, cheap and portable. Its detection is high effective, accurate, and convenient. It can be used for on-site monitoring and is suitable for qualitative and quantitative screenings of a great number of samples. And thus, it will play an important role in the detection of dinitolmide.