Résumé :
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Introduction The ATPase properties of different muscle fiber types is widely used for histochemical diagnostic purposes. The ATPase staining method has a central position in most routine muscle biopsy diagnostic laboratories. However, the method is very laborious and there are disadvantages such as weakening of staining over time and the non-specific staining of capillaries making a distinction of highly atrophic muscle fibers difficult. Since fiber types by ATPase are paralleled by differences in myosin heavy chain isoforms the full assessment of fiber type distribution can also be achieved by immunohistochemical staining. Objectives To develop a reliable and advanced immunohistochemical method for the detection and separation of different fiber types and their subtypes on one slide only. Methods Frozen sections from muscle biopsies were used for myosin double stainings made with the BondMax (Vision biosystems) and/or BenchMark (Ventana medicals) immuno-stainers, using antibodies against myosin heavy chains: slow myosin and myosin A4.74. Slow myosin was stained using a DAB based detection kit with hematoxylin counterstaining and myosin A4.74 with an AP red based detection system. Histochemical ATPase stainings were also performed on serial sections of the same normal controls and myotonic dystrophy type2 samples to compare the fiber type distribution. Results All ATPase based fiber types were easily separated by the double immunostaining technique. Slow type1 fibers stained brown, fast type 2A fibers stained with a deep shade of pink and type 2C fibers being myosin isoform hybrid fibers stained reddish-brown. Moreover, this technique was able to also separate type IIx fibers with a light shade of pink not accessible by conventional ATPase method. Capillaries or other structures were not immunostained. Discussion The immunohistochemical double staining of myosin heavy chains seems to be a reliable method for the detection and separation of different fiber types and subtypes. Immunohistochemical double staining of myosins can be used as an alternative for the more laborious ATPase staining method in routine histopathology, and proved to provide even more detailed information of fast subtypes and highly atrophic fibers. immunostaining technique.
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