PurposeWe aimed to (i) investigate differences in 1-minute sit-to-stand test (1MSTST) performance (i.e., the number of repetitions) between a standardised modality (i.e., starting from a conventional chair with 46 cm seat height) and an individualised modality (i.e., starting with a knee joint flexion angle of 90°), and to (ii) quantify the influence of tibia and femur length on 1MSTST performance.MethodsHealthy participants were recruited for this randomised crossover study, performing each 1MSTST modality twice in a randomised order. The primary outcome was the number of repetitions in the 1MSTST. Secondary endpoints were the acute responses in peripheral oxygen saturation, heart rate, and leg fatigue and dyspnoea. Additionally, we investigated correlations of performance with knee extensor strength in both modalities.ResultsThirty participants were recruited and completed the study. They achieved significantly less repetitions in the standardised 1MSTST compared to the individualised 1MSTST (B = − 12.1, 95% confidence interval [95% CI] =  − 14.8/− 9.4, p < 0.001). We found a significant effect of femur length on 1MSTST performance (B = − 1.6, 95% CI = − 2.6/− 0.7, p = 0.01), tibia length showed significant interaction with the 1MSTST modality (B = 1.2, 95% CI = 0.2/2.2, p = 0.03).ConclusionAn individualisation of the 1MSTST starting position to 90° knee flexion angle leads to more repetitions compared to the traditional starting position. The higher repetition count is explained by controlling for differences in tibia length. We recommend individualisation of the 1MSTST, enabling more valid comparisons across populations and study samples.Trial registration numberhttp://www.ClinicalTrials.gov, NCT04772417.Trial registration dateFebruary 26, 2021.