The invention discloses a tumor chemotherapy drug susceptibility detection kit which comprises a histocyte dispase, an antibiotic solution, an amphotericin solution, an EGTA-Trypsin (Ethylene Glycol Tetraacetic Acid) solution, a serum-containing culture medium DF10, a serum-free culture medium Kertin-SFM, a specimen preserving fluid, a specimen cleaning fluid, a gel coating culturing bottle, a cell fixing fluid, cell dispase, a cell filtering membrane, a neutral red dyeing solution and a collagen gel preparing reagent. A detection operation flow of the kit comprises the steps: primary cell culture, culture in collagen gel drops, and analysis of chemotherapy drug susceptibility. Compared with the prior art, the tumor chemotherapy drug susceptibility detection kit has the beneficial effects that the tumor chemotherapy drug susceptibility detection kit is capable of maximally protecting activity of tumor cells and shortening the drug susceptibility detection time. The tumor chemotherapy drug susceptibility detection kit contains all reagents required for tumor cell primary culture and chemotherapy drug detection; consumable items are disposable articles, thus bacterium pollution in an experiment process is reduced, and the convenience is brought for the use.