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The aim of this study was to develop and validate an automated accelerometry-based system for estimating initial contact (IC) and terminal contact (TC) timing information from walking patterns of healthy control subjects and transtibial amputees that can be used in daily life with minimal interference of researchers. Subjects were instrumented with two uniaxial accelerometers just below the knee while synchronized ground reaction force (GRF) recordings were used as reference measurements. An automated multiphase algorithm was developed to estimate the time of IC and TC in the acceleration signals of five healthy subjects and two transtibial amputees walking at different walking speeds. The accuracy of the detection algorithm in ten control subjects and eight transtibial amputees indicated mean errors ranging between 0.013 and 0.034 s for the TC and IC timing, with 95% confidence interval of the individual step errors ranging between -0.062 and 0.115 s. Correlation coefficients between the estimated stance phase duration and GRF data were 0.98 and 0.97 for controls and amputees, respectively. We concluded that IC and TC can be accurately and easily measured using this system in both healthy subjects and transtibial amputees walking at different walking speeds. The system can be used in clinical situations or gait labs as well as during daily life.