The aim was to determine whether eccentric strengthening changed the muscle architecture of human biceps femoris and consequently, knee range of motion. Twenty-two subjects were randomly assigned to control and experimental groups. The experimental group completed an eccentric strengthening programme for 8 weeks. Outcome measures included hamstring muscle strength (one repetition maximum), the passive knee extension test (PKE) (knee joint angle at which the onset of passive tension occurs), fascicle length (FL) and pennation angle (PA). One repetition maximum increased by 34% (P < 0.01), the PKE test revealed a 5% increase in joint range of motion (P = 0.01), FL increased by 34% (P = 0.01) and PA did not change (P = 0.38). This is the first report of an increase in FL in the biceps femoris following eccentric resistance training. In addition, the results might imply that this fascicle lengthening could lead to an increase in the range of motion of the knee. Clinical implications for rehabilitation and injury prevention are discussed.