Subgroups of fast twitch (FT) muscle fibres were identified by histochemical techniques on muscle samples of m. quadriceps femoris from six male and six female subjects, who had been assigned to three groups; untrained, active and well trained (endurance runners). Slow twitch (ST) and FT fibres were initially identified using the histochemical stain for myofibrillar ATPase, preincubated at pH 10.3 and 4.3. Three people, working independently, then identified the subgroups FTa and FTb on the basis of the staining intensity for only one of the following reactions: α-glycerophosphate dehydrogenase, α-GPD; NADH tetrazolium reductase, NADH-TR; and myofibrillar ATPase preincubated at pH 4.6, ATPase (4.6). FTa fibres were clearly distinguished from the darker staining FTb fibres using the ATPase (4.6) reaction. Differences in the staining within the FT fibres using the α-GPD and NADH-TR reactions were more subtle, and differences between subject groups were evident. The percentage of FTa fibres was overestimated for the untrained and underestimated for the well trained subjects using NADH-TR. With the α-GPD stain the percentage of FTa fibres was generally underestimated. When the data for all three stains were compared, only 27% of the FT fibres were placed in the same subgroups. These results demonstrate that the subgrouping of FT fibres in man is more reliable using the differences in pH sensitivity for the myofibrillar ATPase reaction compared to histochemical reactions for oxidative or glycolytic enzymes.