Demonbreun Alexis R, McNally Elizabeth M
Center for Genetic Medicine, Northwestern University;
Center for Genetic Medicine, Northwestern University.
J Vis Exp. 2015 Dec 23(106):e53551. doi: 10.3791/53551.
Mature muscle has a unique structure that is amenable to live cell imaging. Herein, we describe the experimental protocol for expressing fluorescently labeled proteins in the flexor digitorum brevis (FDB) muscle. Conditions have been optimized to provide a large number of high quality myofibers expressing the electroporated plasmid while minimizing muscle damage. The method employs fluorescent tags on various proteins. Combining this expression method with high resolution confocal microscopy permits live cell imaging, including imaging after laser-induced damage. Fluorescent dyes combined with imaging of fluorescently-tagged proteins provides information regarding the basic structure of muscle and its response to stimuli.
成熟肌肉具有独特的结构，适合进行活细胞成像。在此，我们描述了在拇短屈肌（FDB）中表达荧光标记蛋白的实验方案。已对条件进行了优化，以提供大量表达电穿孔质粒的高质量肌纤维，同时将肌肉损伤降至最低。该方法在各种蛋白质上使用荧光标签。将这种表达方法与高分辨率共聚焦显微镜相结合，可以进行活细胞成像，包括激光诱导损伤后的成像。荧光染料与荧光标记蛋白的成像相结合，可提供有关肌肉基本结构及其对刺激反应的信息。
