Purpose Recently, a fatigue threshold obtained during submaximal repetitive isometric knee extensor contractions was related to VO 2max measured during cycling and to exercise endurance. However, test duration is quite long (20-30 min in young people) to be of practical and possibly clinical use. The purpose of the present study was to test the day-to-day reliability of a newly developed short test that assessed the fatigue threshold during a submaximal test with the knee extensors. Methods Fifteen healthy young males were tested three times, once using the original long protocol (5-min blocks of repetitive unilateral isometric knee extensor contractions with stepwise (5% MVC) increases of force) and twice using a new shorter protocol. In the latter, force increased by 2% MVC every 30 s, starting at 15% MVC (all contractions were 3 s on, 2 s off). The fatigue threshold was defined as the force where the EMG/force ratio started to increase and, compared with the force, at which deoxygenated hemoglobin concentration ([HHb]) increased steeply (HHb threshold). Results The EMG/force threshold during the short trials was reached after 3.9 ± 1.5 min of submaximal exercise and similar (P > 0.05) between days. The EMG/force threshold showed good reliability (intraclass correlation coefficient = 0.87, SEM = 2.2%) and did not differ between (P > 0.05) the short (31.1% ± 7.6% MVC) and long tests (30.5% ± 6.2% MVC), with a significant relation (r = 0.71) between both tests. Similar results (P > 0.05) were found for the HHb threshold. Conclusion In young healthy men, a fatigue threshold can be detected during repetitive isometric knee extensor contractions using a short submaximal test, which may be suitable for untrained or frail people and patients.