In this study the stress protein response to unaccustomed maximal eccentric exercise in humans was investigated. Eleven healthy males performed 300 maximal eccentric actions with the quadriceps muscle. Biopsies from vastus lateralis were collected at 30 minutes, 4, 8, 24, 96, and 168 hours after exercise. Cellular regulation and localization of HSP27, B-crystallin and HSP70 were analyzed by immunohistochemistry, ELISA technique and western blotting. Additionally, mRNA levels of HSP27, B-crystallin and HSP70 were quantified by Northern blotting. Thirty minutes after exercise 81±8% of the myofibers showed strong HSP27 staining (p<0.01) that gradually decreased during the following week. B-crystallin mimicked the changes observed in HSP27. Thirty minutes after exercise the ELISA analysis showed a 49±13% reduction of the HSP27 level in the cytosolic fraction (p<0.01), while western blotting revealed a 15-fold increase of the HSP27 level in the myofibrillar fraction (p<0.01). The cytosolic HSP70 level increased to 203±37% of control level 24 hours after exercise (p<0.05). After four days, myofibrillar-bound HSP70 had increased ~10-fold (p<0.01) and was accompanied by strong staining on cross sections. mRNA levels of HSP27, B-crystallin and HSP70 were all elevated the first day after exercise (p<0.01); HSP70 mRNA showed the largest increase (20-fold at eight hours). HSP27 and B-crystallin seemed to respond immediately to maximal exercise by binding to cytoskeletal/myofibrillar proteins, probably to function as stabilizers of disrupted myofibrillar structures. Later, mRNA and total HSP protein levels, especially HSP70, increased, indicating that HSPs play a role in skeletal muscle recovery and remodeling/adaptation processes to high-force exercise.