The invention discloses a detection kit based on Trinder reaction and application of the detection kit. The invention belongs to the technical field of clinical chemical detection. The kit consists ofa reagent R1 and a reagent R2, wherein the R1 reagent and the R2 reagent contain enzymes for catalyzing a to-be-detected object to generate hydrogen peroxide, the reagent R1 further contains a buffersolution, a preservative and peroxidase, the reagent R2 contains a buffer solution, a preservative, peroxide and a chromogen substrate, the concentration of peroxidase in the reagent R1 is 100-150 KU/L, and the reagent R1 further contains 4-aminoantipyrine with the concentration of 0.5-1 g/L. the peroxidase and the 4-aminoantipyrine in the reagent R1 can be used for removing clinical drugs capable of participating in the Trinder reaction in a to-be-detected object before the Trinder reaction, so that a detection result is more accurate. According to the invention, other substances do not needto be additionally added into a reaction system, and other impurities cannot be introduced; the method is simple and efficient, can be widely applied to various clinical chemical detections, avoids the interference of clinical drugs, enhances the accuracy of detection results, and also improves the functional sensitivity of the kit.