Abstract: Objective：To explore and analyze the influencing factors of delayed elimination combined with acute kidney injury after high-dose methotrexate (HDMTX) chemotherapy on a case of children with acute lymphoblastic leukemia (ALL)， so as to promote rational clinical medication. Methods： 1 case of children with ALL occured delayed elimination of MTX combined with acute kidney injury in the course of HDMTX chemotherapy. By consulting the hospital electronic medical record system， collecting information about the diagnosis， treatment， examination and medication of the pationt， using fluorescence in situ hybridization for genotyping of methotrexate related gene mutation sites， and analyzed the related factors that may cause MTX excretion abnormalities. Results：The delayed elimination and acute kidney injury in the course of HDMTX chemotherapy may be related to the amount of hydration urine alkalization， drug metabolism enzymes and other factors. Conclusion： The genotype of MTX should be detected before HDMTX chemotherapy in ALL children， and individualized treatment plan should be made according to the genotype of patients， and MTX blood concentrations， liver and kidney functions of the patients and other factors affecting MTX excretion should be timely monitored during chemotherapy， so as to improve the efficacy and reduce the occurrence of toxic and side effects of MTX.