Transmission Electron Microscopy to Quantify Glycogen Distribution in Human Skeletal Muscles
Published: July 31, 2023
Abstract
Source: Jensen, R., et al. Quantification of Subcellular Glycogen Distribution in Skeletal Muscle Fibers using Transmission Electron Microscopy. J. Vis. Exp. (2022)
In this video, we demonstrate the sample preparation of human skeletal muscle tissue and its staining for transmission electron microscopy to visualize and quantify subcellular glycogen distribution.
Protocol
All procedures involving human participants have been performed in compliance with the institutional, national, and international guidelines for human welfare and have been reviewed by the local institutional review board 1. Primary fixation, post-fixation, embedding, sectioning, and contrasting Prepare 1.6 mL of primary fixative solution (2.5% glutaraldehyde in 0.1 M sodium cacodylate buffer (pH 7.3)) in a 2 mL micro centrifugation tube. Store it at 5 °C for …
Representative Results
Figure 1: Imaging and fiber typing. (A) Each fiber is imaged in a randomized systematic order. (B) Example of an image from the subsarcolemmal space. (C) Example of an image from the myofibrillar space. (D) In each myofibrillar image, the width of one Z-disc is measured (red lines). The measurements of a total of 12 Z-discs (one per image) give …
Açıklamalar
The authors have nothing to disclose.
Materials
| 1,2-Propylene oxide | Merck | 75-56-9 | |
| Embedding 812 resin medium kit | Taab | T031 | |
| Glutaraldehyde solution 25% | Merck | 1.04239.0250 | |
| ITEM | Olympus | Imaging software | |
| Leica EM AC20 | Leica | Automatic contrasting system | |
| OSIS Veleta digital camera | Olympus | ||
| Osmium tetroxide 4% solution | Polysciences | 0972A | |
| Philips CM 100 Transmission EM | Philips | ||
| Potassium hexacyanoferrate (II) trihydrate | Sigma-Aldrich | 455989-245G | |
| Sodium cacodylatbuffer 0.2 M ph 7.4 | Ampliqon.com | AMPQ40989.0500 | |
| Ultra-microtome Leica UC7 | Leica | ||
| Ultrostain lead citrate 3%, stabilized solution | Leica | 16707235 | |
| Uranyl acetate dihydrate | Polysciences | 6159-44-0 |
Etiketler
Bu Makaleden Alıntı Yapın
Transmission Electron Microscopy to Quantify Glycogen Distribution in Human Skeletal Muscles. J. Vis. Exp. (Pending Publication), e21462, doi: (2023).