Residual force enhancement (FE) following stretch of an activated muscle is a well accepted property of skeletal muscle contraction. However, the mechanism underlying FE remains unknown. A crucial assumption on which some proposed mechanisms are based is the idea that forces in the enhanced state cannot exceed the steady-state isometric force at a sarcomere length associated with optimal myofilament overlap. Although there are a number of studies in which forces in the enhanced state were compared to the corresponding isometric forces on the plateau of the force-length relationship, these studies either did not show enhanced forces above the plateau, or if they did, they lacked measurements of sarcomere lengths confirming the plateau region. Here, we revisited this question by optimizing stretch conditions and measuring the average sarcomere lengths in isolated fibers, and found that FE exceeded the maximal isometric reference force obtained at the plateau of the force-length relationship consistently (mean±SD: 4.8±2.1%) and by up to 10%. When subtracting the passive component of FE from the total FE, the enhanced forces remained greater than the isometric plateau force (mean±SD: 4.3±2.0%). Calcium-induced increases in passive forces, known to be present in single fibers and myofibrils, are too small to account for the FE observed here. We conclude that FE cannot be explained exclusively with stretch-induced development of sarcomere length non-uniformities, that FE in single fibers may be associated with the recruitment of additional contractile force, and that isometric steady-state forces in the enhanced state are not uniquely determined by sarcomere lengths.