Methodology Report

Adherent Primary Cultures of Mouse Intercostal Muscle Fibers for Isolated Fiber Studies

Figure 6

Overexpression of foreign proteins in intercostal fibers in culture. FDB (a, b, e, f) and intercostal (c, d, g, h) muscle fibers expressing either GFP (a–d) or a Glut4-GFP fusion construct (e–h). Scale bars are 50 μm (b, d) or 25 μm (f, h). Images were acquired as in Figure 4 for panels (a–d) or with an Olympus FluoView 500 confocal system, built on an Olympus IX71 inverted microscope for panels (e–h). Fibers were imaged viewed with a UPlan Apo 60x/NA 1.20 water-immersion objective lens using 488 nm excitation (Multiline Argon laser) for GFP with emission detected with 505 nm long-pass filter. Photobleaching and photodamage were reduced by operating the lasers at the lowest possible power that still provided images with satisfactory signal to noise ratio. Brightfield images in panels (b, d, f, h) were adjusted for improved contrast using ImageJ.
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