A One-Step Immunostaining Method to Visualize Rodent Muscle Fiber Type within a Single Specimen.

A One-Step Immunostaining Method to Visualize Rodent Muscle Fiber Type within a Single Specimen.
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DOI:
10.1371/journal.pone.0166080
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发表时间:
2016
期刊:
影响因子:
3.7
通讯作者:
Mizunoya W
Mizunoya W
中科院分区:
综合性期刊3区
文献类型:
--
作者:
Sawano S;Komiya Y;Ichitsubo R;Ohkawa Y;Nakamura M;Tatsumi R;Ikeuchi Y;Mizunoya W

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在这项研究中,我们提出了一种四重免疫染色方法,用于快速肌纤维分型的小鼠和大鼠使用特异性抗体的成人肌球蛋白重链(MyHC)亚型MyHC1,2A,2X和2B,这是常见的标记蛋白的不同的肌纤维类型。我们开发了大鼠单克隆抗体特异性的每一个MyHC亚型和共轭这四个抗体的荧光团具有不同的激发和发射波长。通过混合四种类型的缀合抗体,MyHC1、2A、2X和2B可以在单个标本中区分,从而允许容易地描绘骨骼肌纤维类型。此外,我们可以在小鼠和大鼠的单个纵向肌肉切片中观察到同时表达MyHC2X和MyHC2B的杂交纤维,这在以前的技术中是无法实现的。这种染色方法有望应用于研究肌纤维类型转变对环境因素的响应,并最终开发出调控动物肌纤维类型的技术。
In this study, we present a quadruple immunostaining method for rapid muscle fiber typing of mice and rats using antibodies specific to the adult myosin heavy chain (MyHC) isoforms MyHC1, 2A, 2X, and 2B, which are common marker proteins of distinct muscle fiber types. We developed rat monoclonal antibodies specific to each MyHC isoform and conjugated these four antibodies to fluorophores with distinct excitation and emission wavelengths. By mixing the four types of conjugated antibodies, MyHC1, 2A, 2X, and 2B could be distinguished within a single specimen allowing for facile delineation of skeletal muscle fiber types. Furthermore, we could observe hybrid fibers expressing MyHC2X and MyHC2B together in single longitudinal muscle sections from mice and rats, that was not attained in previous techniques. This staining method is expected to be applied to study muscle fiber type transition in response to environmental factors, and to ultimately develop techniques to regulate animal muscle fiber types.
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