The instrumented Timed "Up and Go" test (iTUG) has the potential for playing an important role in providing clinically useful information regarding an individual's balance and mobility that cannot be derived from the original single-outcome Timed "Up and Go" test protocol. The purpose of this study was to determine the reliability and validity of the iTUG using body-fixed inertial sensors in people affected by stroke. For test-retest reliability analysis, 14 individuals with stroke and 25 nondisabled elderly patients were assessed. For validity analysis, an age-matched comparison of 12 patients with stroke and 12 nondisabled controls was performed. Out of the 14 computed iTUG metrics, the majority showed excellent test-retest reliability expressed by high intraclass correlation coefficients (range 0.431-0.994) together with low standard error of measurement and smallest detectable difference values. Bland-Altman plots demonstrated good agreement between two repeated measurements. Significant differences between patients with stroke and nondisabled controls were found in 9 of 14 iTUG parameters analyzed. Consequently, these results warrant the future application of the inertial sensor-based iTUG for the assessment of physical deficits poststroke in longitudinal study designs.