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This study aimed to develop new prediction models from directly determined peak of oxygen uptake (VO 2peak ) in adolescents using 20-metre shuttle run test (20 m-SR) and to compare the new models with previously published equations. This study included 148 adolescents (43% girls), aged 13.37 ± 1.84 years old. Adolescents were randomly assigned to validation (n = 91) and cross-validation (n = 57) groups. VO 2peak was measured using a gas analyser in both maximal exercise tests in the laboratory as well as by 20 m-SR. The multiple linear regression method was applied to develop the models using BMI, BMI-z score and body fat percentage (%FM). The proposed models presented better predictive performance (R2 = 75.6% to 78.9%) and a lower absolute percentage error (%error = 10.51 to 11.78%) than the previously published equations (R2 = 38.8% to 69.1%; %error = 13.13% to 21.54%). The Model with BMI-z was the best fit equation in girls, and the model with BMI-z and %FM in boys. Therefore, it is recommended that the equations developed in the present study be used in future research and projects in the school environment to estimate VO 2peak in adolescents by the 20 m-SR test.