Quantification of Subcellular Glycogen Distribution in Skeletal Muscle Fibers using Transmission Electron Microscopy
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Quantification of Subcellular Glycogen Distribution in Skeletal Muscle Fibers using Transmission Electron Microscopy. / Jensen, Rasmus; Ørtenblad, Niels; Di Benedetto, Cristiano; Qvortrup, Klaus; Nielsen, Joachim.
I: Journal of Visualized Experiments, Bind 2022, Nr. 180, e63347, 2022.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
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TY - JOUR
T1 - Quantification of Subcellular Glycogen Distribution in Skeletal Muscle Fibers using Transmission Electron Microscopy
AU - Jensen, Rasmus
AU - Ørtenblad, Niels
AU - Di Benedetto, Cristiano
AU - Qvortrup, Klaus
AU - Nielsen, Joachim
N1 - Publisher Copyright: © 2022 JoVE Journal of Visualized Experiments.
PY - 2022
Y1 - 2022
N2 - With the use of transmission electron microscopy, high-resolution images of fixed samples containing individual muscle fibers can be obtained. This enables quantifications of ultrastructural aspects such as volume fractions, surface area to volume ratios, morphometry, and physical contact sites of different subcellular structures. In the 1970s, a protocol for enhanced staining of glycogen in cells was developed and paved the way for a string of studies on the subcellular localization of glycogen and glycogen particle size using transmission electron microscopy. While most analyses interpret glycogen as if it is homogeneously distributed within the muscle fibers, providing only a single value (e.g., an average concentration), transmission electron microscopy has revealed that glycogen is stored as discrete glycogen particles located in distinct subcellular compartments. Here, the step-by-step protocol from tissue collection to the quantitative determination of the volume fraction and particle diameter of glycogen in the distinct subcellular compartments of individual skeletal muscle fibers is described. Considerations on how to 1) collect and stain tissue specimens, 2) perform image analyses and data handling, 3) evaluate the precision of estimates, 4) discriminate between muscle fiber types, and 5) methodological pitfalls and limitations are included.
AB - With the use of transmission electron microscopy, high-resolution images of fixed samples containing individual muscle fibers can be obtained. This enables quantifications of ultrastructural aspects such as volume fractions, surface area to volume ratios, morphometry, and physical contact sites of different subcellular structures. In the 1970s, a protocol for enhanced staining of glycogen in cells was developed and paved the way for a string of studies on the subcellular localization of glycogen and glycogen particle size using transmission electron microscopy. While most analyses interpret glycogen as if it is homogeneously distributed within the muscle fibers, providing only a single value (e.g., an average concentration), transmission electron microscopy has revealed that glycogen is stored as discrete glycogen particles located in distinct subcellular compartments. Here, the step-by-step protocol from tissue collection to the quantitative determination of the volume fraction and particle diameter of glycogen in the distinct subcellular compartments of individual skeletal muscle fibers is described. Considerations on how to 1) collect and stain tissue specimens, 2) perform image analyses and data handling, 3) evaluate the precision of estimates, 4) discriminate between muscle fiber types, and 5) methodological pitfalls and limitations are included.
U2 - 10.3791/63347
DO - 10.3791/63347
M3 - Journal article
C2 - 35188121
AN - SCOPUS:85125004595
VL - 2022
JO - Journal of Visualized Experiments
JF - Journal of Visualized Experiments
SN - 1940-087X
IS - 180
M1 - e63347
ER -
ID: 300757859