Approximately 15-20% of paediatric acute lymphoblastic leukaemia (ALL) patients currently fail treatment. Since the number of agents for testing outweighs patient availability for clinical trials, robust preclinical models are required to prioritise effective drugs. Patient-derived xenografts (PDXs) of ALL in immune-deficient mice provide a clinically relevant testing model enabling monitoring of engraftment and drug response by flow cytometric enumeration of % human vs mouse CD45+ cells in mouse peripheral blood (PB). However, this model overlooks leukaemic harbour / relapse sites, as well as organ-specific drug efficacy. This study introduced a bioluminescence imaging (BLI) capability into paediatric ALL PDXs to enhance monitoring of leukaemia burden. BLI and PB were used to track engraftment and response to an induction-type drug regimen; vincristine (0.15mg/kg weekly x4), dexamethasone (5mg/kg daily x5 x4) and L-asparaginase (1,000KU/kg daily x5 x4) (VXL). Additional groups received 0.5, 0.25 and 0.125x this dose for 2 weeks. Both measures correlated strongly (R2=0.8, p=<0.0001) though BLI detected engraftment 6 weeks before PB. VXL response was scored using clinically relevant criteria; the 0.125, 0.25, 0.5 and 1x groups scored Progressive Disease 2, Stable Disease, Complete Response (CR) and Maintained CR, respectively. BLI measured regression with greater range and sensitivity than PB, highlighting differences between the 1x and 0.5x cohorts. Treatment caused a 3.2-fold decrease in bioluminescence in the 0.5x group, compared to >2,000-fold decrease in the 1x group. Day 28 culls revealed <1% blasts in the bone marrow (BM) in the 1x group, yet >30% in the 0.5x group, well above the clinical definition of a CR of 5%. By BLI, treatment caused >2000-fold decrease in BM bioluminescence in the 1x group compared to a 1.3-fold increase in the 0.5x group. These results show that BLI enhances sensitivity of preclinical drug efficacy testing, allowing for more accurate drug prioritisation for clinical trials.