The collection of muscle samples in horses to estimate their performance potential, adaptations to training and diagnosis of exertional myopathies is of increasing interest. This study was designed to examine whether repeated biopsying of the gluteus medius muscle at 7 week intervals alters morphology and muscle fibre variables in the sampling area. The gluteus had been biopsied 3 times previously, each biopsy separated by a 7 week interval. Seven weeks after the last biopsy, this study was initiated. Percutaneous needle biopsies were taken from left and right muscles at 2 depths (2 and 6 cm) and at 2 sites for each depth (22 and 25 cm from the tuber coxae) of 7 mature Thoroughbreds. The 22 cm site was located close (2 cm caudal) to an area of the muscle that had been biopsied previously. The 25 cm site was 5 cm apart from this area. A total of 52 samples were available for histology, electron microscopy and immunohistochemistry. The latter was used to study percentages, sizes and capillaries of muscle fibre types. Most muscle samples collected (75%) had normal morphology, but the remaining 25% showed signs of muscle repair. In some circumstances, they showed regenerative signs of complete muscle repair, while in others ineffective muscle repair (scar formation) was evident. Nevertheless, sufficient normal tissue was always available for measuring routine muscle variables. Samples collected 3 cm laterally apart showed large differences with regard to muscle fibre type variables examined. These results show that repeated muscle samples in intervals of 7 weeks do not have effects that would impair evaluation of muscle fibre variables for diagnosis of effectivity of conditioning programmes. Furthermore, they emphasise that the only way of keeping variations through sample collection technique small is to standardise all biopsying techniques accurately.